JP2013028546A - Agent for increasing serotonin in brain - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a composition which exerts an effect that increases serotonin in the brain through oral ingestion.SOLUTION: The agent for increasing serotonin in the brain contains an epicatechin obtained by steaming and drying camellia leaves to obtain dried camellia leaves, extracting the obtained dried camellia leaves with distilled water or ethanol, supplying the extract to an ion exchange resin column, and concentrating and drying the effluent by 20% ethanol.

Description

  The present invention relates to an agent that exerts an increase in serotonin in the brain and a food or drink containing the same.

  Serotonin (5-hydroxytryptamine; 5-HT) is a kind of physiologically active amine and belongs to indoleamines. There are about 10 mg in the human body, 2% of which exists in the central nervous system and is known as an important neurotransmitter. Central serotonergic pathways that project extensively from the brainstem, such as the raphe nucleus, to the cerebrum and spinal cord, regulate emotion and mood, sleep and wakefulness, appetite, exercise, body temperature, and cognition through its many receptor subtypes. Involved (Non-Patent Documents 1 and 2).

The neurotransmitter serotonin in the brain is a substance related to mental stability, and an increase in 5-HT has an effect of reducing it against stress. Particularly related to stress are 5-HT, γ-aminobutyric acid (GABA), dopamine (DA), and noradrenaline (NA) (Non-patent Document 3). When 5-HT is insufficient, mood becomes unstable. Become mentally unstable, such as falling asleep or waking up. It is generally known that 5-HT increases with awakening, and when 5-HT increases, the brain is activated and becomes clean, mentally positive and calm. Reference 4).
For example, when depression occurs, NA and 5-HT deficiency occurs in central monoamine neurons related to emotions and sleep in the brain, resulting in inadequate chemical transmission and depressive symptoms on both mind and body (Non-Patent Document 5). In particular, 5-HT is considered to be the main factor involved in depression (Non-Patent Document 6), and it is also known that 5-HT concentration is decreased in the brain of suicides (Non-Patent Document 6). 7). Therefore, if 5-HT can be increased, it is thought that the refreshing feeling in the morning and daytime will be improved, the mood will be calmed down, stress will be reduced, and depression will be improved.

  Examples of materials that have been reported to increase serotonin in the brain, such as serotonin release promoting action and serotonin reabsorption inhibiting action, include kale, propolis or an extract thereof (Patent Document 11), N-anisoyyl-γ-amino. There are butyric acid or p-anisic acid (patent document 12), glycine (non-patent document 9), St. John's wort (non-patent document 10) and the like, and there are selective serotonin reuptake inhibitors (SSRI) as drugs. There may be side effects such as loss of appetite, weight gain or loss, and abnormal libido, and a stable and safe material is required as one of the options for increasing serotonin.

  Epicatechin, that is, 3,3 ′, 4 ′, 5,7-pentahydroxyflavan is one of the flavonoids contained in catechins, widely contained in the skin and trunk of the resin, and particularly in abundant amounts in Gambia and areca It is. In addition, the carbons at the 2nd and 3rd positions are asymmetric carbons in terms of structure, and can have four types of structures even in the same chemical structure. These are distinguished from (−)-catechin, (+)-catechin, (−)-epicatechin, (+)-epicatechin. Epicatechin here is (−)-epicatechin and is known to have various physiological actions. For example, pancreatic β-cell protective agent (Patent Document 1), method and composition for improving cognitive function (Patent Document 2), hyaluronidase activity inhibitor (Patent Document 3), antioxidant and method for producing the same, and Food / beverage products containing antioxidants (Patent Document 4), skin whitening external preparation (Patent Document 5), bifidobacteria growth promoter (Patent Document 6), heat-treated cocoa products useful for improving vascular health ( Patent Document 7), Adiponectin secretion promoting composition, food and drink containing the composition (Patent Document 8), deodorant composition (Patent Document 9), tryptase activity inhibitor and use thereof (Patent Document 10), etc. It is disclosed.

Regarding brain or 5-HT, in the brain perfusion test using the microdialysis method, catechins have an action to increase 5-HT release in the rat brain hippocampus. In particular, epicatechin has epicatechin gallate and (+ ) -Catechin and other catechins are known to have an increase effect of about 100 times or more. Moreover, about catechin, the release of 5-HT was seen also by oral administration (nonpatent literature 8). It has also been reported that epicatechin has an inhibitory effect on serotonin uptake that contributes to serotonin regulation in a test system in which synaptosomes are isolated and cultured (Non-patent Document 11). However, these studies did not evaluate the case where epicatechin was orally ingested, and even when epicatechin was ingested, it was sulfate-conjugated and glucuronidated by intestinal absorption and did not migrate to the brain as epicatechin itself. Therefore, from the results of Non-Patent Document 11, it cannot be determined whether or not the conjugate has an action on serotonin.
Examples of raw materials containing epicatechin include green tea leaves, cacao beans, grape seeds, apple peels, soybeans, broad beans, camellia leaves, and the like.

JP 2008-7452 A Special table 2009-539991 Japanese Patent Laid-Open No. 06-9391 JP 2007-254508 A JP-A-10-120546 Japanese Patent Application Laid-Open No. 2006-288221 JP-T 2009-501161 JP 2006-131512 A JP 09-38183 A JP 2007-186457 A Japanese Patent Laid-Open No. 2003-300892 JP 2002-154958 A

Annuals New York Academy of Science, 600, 9-35, 1990 Pharmacology Biochemistry and Behavior, 54, 129-141, 1996 "New Bioscience Series: Exploring Stress-When Seen at the Molecular Level", Science Doujin, Kyoto, 81-91, 1994 Serotonin-deficient brain-reworking crisp and blue brains-Japan Broadcasting Publishing Association, Tokyo, 49-56, 2003 "Depression Science and Health-For General Physicians", Asakura Shoten, Tokyo, 15-20, 1987 Journal of Pharmacy and Pharmacology, 57, 651-656, 2005 The British Journal of Psychiatry, 113 (505), 1407-1411, 1967 Abstracts of Japanese Society of Nutrition Foods, 2D-13p, 2008 Psychiatry and Clinical Neurosciences, 65, 142-149, 2011 British Journal of Pharmacology, 142, 414-418, 2004 Phytomedicine, 14, 396-402, 2007

  Providing a composition that exerts an increasing effect on intracerebral serotonin when taken orally.

As described above, in the prior art, no study has been made on the case where epicatechin is taken orally. Therefore, the present invention examined in detail whether or not serotonin in the brain is increased even when epicatechin is orally ingested. As a result, it was confirmed that serotonin in the brain was increased only by ingesting epicatechin intragastrically into rats by single oral intake. Moreover, camellia leaf was found as a raw material of epicatechin, and it was confirmed that serotonin in the brain increased in a composition in which epicatechin in the extract was highly concentrated. In other words, the solution of the present invention is to exert a serotonin-increasing action in the brain by orally ingesting an epicatechin-containing material, particularly a camellia leaf extract.
That is, an object of the present invention is to increase the amount of serotonin in the brain, and as a result of research, it has been found that serotonin in the brain increases only by taking epicatechin once by intragastric administration. Furthermore, a method for preparing a composition containing only epicatechin and not containing other catechins from camellia leaves was established, and an increasing action on brain serotonin was found.

  In the oral intake of epicatechin, the serotonin in the brain was effectively increased, and a method for preparing a composition containing only epicatechin in high concentration from camellia leaves and not containing other catechins was established.

It is a graph which shows the 5-HT discharge | release amount by epicatechin administration. It is a graph which shows the 5-HT discharge | release amount by catechin mixture administration.

  The present invention relates to a brain serotonin increasing agent containing epicatechin.

  The present invention also relates to a brain serotonin increasing agent containing a fraction containing high epicatechin, wherein epicatechin is extracted from camellia leaves.

  Furthermore, the present invention is a method for preparing epicatechin from camellia leaves, the camellia leaves are steamed and dried to obtain dried camellia leaves, and the obtained dried camellia leaf distilled water or ethanol extract is used as an ion exchange resin. The present invention relates to a method characterized in that a fraction containing high epicatechin is obtained by applying to a column and concentrating and drying a 20% ethanol eluate.

  Furthermore, this invention relates to the foodstuff containing the serotonin increasing agent in a brain containing epicatechin.

  The present invention relates to a composition that exhibits an effect of increasing brain serotonin levels when taken orally. Serotonin is a neurotransmitter in the brain that is related to mood and arousal, and is generally said to refresh the mood and increase concentration. Therefore, a composition that raises the serotonin level has the possibility of imparting effects such as mood change and concentration enhancement, and can be expected to be applied as a product as a functional material for confectionery and health foods.

  Moreover, this invention provides the food / beverage products containing the serotonin increasing agent in a brain containing the said epicatechin. Foods and drinks include fresh foods, animal foods such as meat and fish, vegetable foods such as cereals and vegetables, processed foods such as dairy products, bread and instant foods, taste foods such as confectionery, sweeteners and seasonings Ingredients for cooking seasonings such as, health foods, special-purpose foods, water, soft drinks, alcoholic beverages, beverages such as tea, food processing materials, food additives and the like.

[Example 1]
Test of the effect of epicatechin on the release of serotonin in the brain

(Test animal)
In this test, Wistar male rats (270 to 310 g, 8 to 10 weeks old) were used. Rats were housed on a 12 hour light / dark cycle in an environment of room temperature of about 24 ° C. and humidity of 45-50%. In addition, food and water during breeding were freely consumed.

(Reagents)
Reagents used in this test were (−)-epicatechin [EC (Wako, Osaka), purity 98% or higher] and catechin mixture [Mix (Wako, Osaka), catechins 85% or higher; (−)-epi Gallocatechin gallate (hereinafter referred to as EGCG) approximately 36.2%, (−)-epigallocatechin (hereinafter referred to as EGC) approximately 23.2%, (−)-epicatechin gallate (hereinafter referred to as ECG) approximately 10.6%, EC about 7.4%, (+)-catechin (hereinafter CA) about 2.4%, (-)-gallocatechin about 5.7%, (-)-gallocatechin gallate about 1.6%, etc.] Are adjusted to 1 mg / kg, 10 mg / kg, 50 mg / kg, and 100 mg / kg, respectively, and each reagent is dissolved in 0.5 ml or 1 ml of physiological saline (0.9% NaCl). Directly using an oral sonde It was administered into the rat stomach. As a control, the same amount of physiological saline was used. Here, each administration group used 4-5 rats per group.

(Test method)
Rats were anesthetized with pentobarbital sodium, and a guide cannula was implanted with reference to Paxinos and Watson brain maps. After confirming that the guide cannula was completely fixed, a dummy cannula was inserted. After the rat recovered, it was anesthetized with sodium pentobarbital, a dialysis probe was inserted into the guide cannula, and the dialysate (standard Ringer's solution) was perfused (perfusion rate 1 μl / min). The dialysate was measured for the amount of 5-HT by high performance liquid chromatography every 20 minutes. Sixty minutes later, EC or Mix was administered into the rat stomach and the measurement was continued until the maximum release of 5-HT was observed. All data are expressed as mean (mean) ± standard deviation (SD), and statistical processing is performed using statistical analysis software SPSS17.0J, using Mann-Whitney test (non-paired and non-parametric) and one-way analysis of variance. It was determined that there was a significant difference with a risk rate of less than 5%.

(result)
The results obtained in the brain 5-HT release promoting effect test are shown in FIGS.
As can be seen from FIG. 1, when EC is administered, the amount of 5-HT released is 110 ± 16% at EC 1 mg / kg administration, 133 ± 10% at 10 mg / kg administration, and 513 ± 535% at 50 mg / kg administration. The dose of 100 mg / kg was 987 ± 643%, and the amount of 5-HT released increased in an EC concentration-dependent manner (FIG. 1). There was no significant difference in the amount of 5-HT released after administration of EC 1 mg / kg and 10 mg / kg. In addition, the amount of 5-HT released when the highest concentration of EC 100 mg / kg was administered was significantly increased compared to that when EC 1 mg / kg and 10 mg / kg were administered.
On the other hand, as shown in FIG. 2, when Mix, which is a catechin mixture, was administered, the amount of 5-HT released was 113 ± 12% when Mix 1 mg / kg was administered, 135 ± 17% when administered 10 mg / kg, and 50 mg / kg. It was 148 ± 39% at the dose of kg and 227 ± 112% at the dose of 100 mg / kg, and even in the case of Mix administration, the 5-HT amount tended to increase depending on the concentration.

Compared to the 1 mg / kg and 10 mg / kg doses, 5-HT release was comparable, and both EC and Mix showed a significant increase in 5-HT release from the brain hippocampus at around 10 mg / kg. It turned out not to be a concentration.
Furthermore, when EC 1 mg / kg administration and EC 100 mg / kg administration were compared, there was an approximately 9-fold difference in the amount of 5-HT released. In contrast, in Mix 1 mg / kg administration and Mix 100 mg / kg administration, although there was a 100-fold difference in catechin concentration, the difference in 5-HT release was about 2-fold.
Also, EC 50 mg / kg and 100 mg / kg administration showed an increase in 5-HT release that was about 4-5 times higher than Mix 50 mg / kg and 100 mg / kg administration.
From the above, it was confirmed that serotonin in the brain increased more markedly by epicatechin administration than by catechin mixture administration by oral ingestion.

[Example 2]
Preparation of epicatechin-rich composition from camellia leaves-1
5.477 kg of camellia japonica leaves were steamed on a boiling water bath for 5 minutes and then dried with warm air to obtain 1.89 kg of dried camellia leaves. 18.9 L of distilled water was added to the dried camellia leaf, and the mixture was left standing at room temperature for 4 hours. Insoluble matter was removed by filtration, and the filtrate was applied to a column packed with 3 L of Diaion HP-20 (manufactured by Mitsubishi Chemical Corporation), and eluted sequentially with 6 L of distilled water, 6 L of 10% ethanol, and 6 L of 20% ethanol. . The 20% ethanol eluate was concentrated under reduced pressure and freeze-dried to obtain 23.52 g of an epicatechin-rich fraction (epicatechin content 24.7%).

[Example 3]
Preparation of epicatechin-rich composition from camellia leaves-2
151.64 g of camellia leaves were steamed on a boiling water bath for 5 minutes and then dried with warm air to obtain 58.48 g of dried camellia leaves. 1.175 L of distilled water was added to the dried camellia leaf, and the mixture was left standing at room temperature for extraction overnight. The insoluble material was removed by filtration, and the filtrate was freeze-dried to obtain 12.18 g of camellia leaf water extract. The epicatechin content of the camellia leaf water extract was 3.12%. 10 g of the resulting camellia leaf water extract was dissolved in 1 L of distilled water and applied to a column packed with 250 mL of Diaion HP-20 (manufactured by Mitsubishi Chemical Corporation). 1 L of distilled water, 1 L of 10% ethanol, 1 L Of 20% ethanol. The 20% ethanol eluate was concentrated under reduced pressure and freeze-dried to obtain 0.932 g of an epicatechin-rich fraction (epicatechin content 32.94%).

[Example 4]
Preparation of epicatechin-rich composition from camellia leaves-3
Camellia leaves 90.12 g were steamed on a boiling water bath for 5 minutes and then dried with warm air to obtain 34.50 g of dried camellia leaves. 690 mL of 70% ethanol was added to the dried camellia leaf, and the mixture was extracted by standing at room temperature overnight. The insoluble material was removed by filtration, and the filtrate was concentrated under reduced pressure and lyophilized to obtain 6.58 g of a camellia leaf 70% ethanol extract. The epicatechin content of the camellia leaf 70% ethanol extract was 6.75%. 1 g of the resulting camellia leaf 70% ethanol extract was dissolved in 1 L of distilled water, and applied to a column packed with 250 mL of Diaion HP-20 (Mitsubishi Chemical Corporation). 1 L of distilled water, 1 L of 10% ethanol Elute sequentially with 1 L of 20% ethanol. The 20% ethanol eluate was concentrated under reduced pressure and freeze-dried to obtain 0.12 g of a high epicatechin-containing fraction (epicatechin content: 44.09%).

[Example 5]
Test for accelerating 5-HT release in brain of fraction containing high epicatechin Using the fraction containing high epicatechin obtained in Example 2 as a sample, the effect on brain 5-HT level was verified in the same manner as in Example 1. Then, it was confirmed that the 5-HT level in the brain was increased by oral administration of the fraction containing high epicatechin.

[Example 6]
Chocolate, troche, chewing gum, candy, gummy jelly, and beverage containing the epicatechin-rich fraction-containing brain serotonin increasing agent of the present invention were produced by conventional methods. Their formulations are shown below. Note that the scope of the present invention is not limited by these.

Chocolate prescription cacao mass 20.0% by weight
Sugar 42.0
Whole milk powder 20.0
Cocoa Butter 17.0
Epicatechin 0.4
Lecithin 0.5
Fragrance 0.1
100.0

Chocolate prescription cacao mass 20.0% by weight
Sugar 41.4
Whole milk powder 20.0
Cocoa Butter 17.0
Epicatechin-rich fraction of Example 2 1.0
Lecithin 0.5
Fragrance 0.1
100.0

Lozenge prescription glucose 73.3% by weight
Lactose 16.0
Gum arabic 6.0
Fragrance 1.0
Sodium monofluorophosphate 0.7
Epicatechin 1.0
Lactose 2.0
100.0

Troche prescription glucose 72.3% by weight
Lactose 16.0
Gum arabic 6.0
Fragrance 1.0
Sodium monofluorophosphate 0.7
Epicatechin-rich fraction of Example 3 2.0
Lactose 2.0
100.0

Chewing gum formula gum base 20.0% by weight
Sugar 54.7
Glucose 15.3
Minamata 9.3
Epicatechin 0.2
Fragrance 0.5
100.0

Chewing gum formula gum base 20.0% by weight
Sugar 54.4
Glucose 15.3
Minamata 9.3
Epicatechin-rich fraction of Example 4 0.5
Fragrance 0.5
100.0

Candy prescription sugar 50.0% by weight
Mizuame 34.0
Citric acid 2.0
Epicatechin 0.6
Fragrance 0.2
Water remaining
100.0

Candy prescription sugar 50.0% by weight
Mizuame 34.0
Citric acid 2.0
Epicatechin term-containing fraction of Example 3 1.0
Fragrance 0.2
Water remaining
100.0

Gummy jelly prescription gelatin 60.0% by weight
Mizuame 20.5
8.5 sugar
Vegetable oil 4.5
Mannitol 3.0
Malic acid 2.0
Epicatechin 1.0
Fragrance 0.5
100.0

Gummy jelly prescription gelatin 59.0% by weight
Mizuame 20.0
8.5 sugar
Vegetable oil 4.5
Mannitol 3.0
Malic acid 2.0
Epicatechin-rich fraction of Example 2 2.5
Fragrance 0.5
100.0

Prescription orange juice 30.0% by weight
Isomerized sugar 15.24
Citric acid 0.10
Vitamin C 0.04
Fragrance 0.10
Epicatechin 0.10
Water remaining
100.0

Prescription orange juice 30.0% by weight
Isomerized sugar 15.24
Citric acid 0.10
Vitamin C 0.04
Fragrance 0.10
The fraction containing high epicatechin content of Example 2 0.50
Water remaining
100.0

  The camellia leaf extract-containing brain serotonin increasing agent can be applied to various products.

Claims (4)

  A serotonin increasing agent in the brain containing epicatechin.   The brain serotonin increasing agent according to claim 1, wherein the epicatechin is a fraction containing high epicatechin content extracted from camellia leaves. A method for preparing a fraction containing high epicatechin content from camellia leaves,
Steamed camellia leaves, dried to obtain dried camellia leaves,
The resulting dried camellia leaf was extracted with distilled water or ethanol and applied to an ion exchange resin column.
A method comprising obtaining a fraction containing high epicatechin content by concentrating and drying a 20% ethanol eluate.   A food containing a serotonin increasing agent in the brain containing epicatechin.

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