WO2005122798A1 - Functional beverage composition for improving ability of learning, concentration and memory - Google Patents

Abstract

The present invention relates to a functional beverage composition for improving concentration, learning ability and memory. Provided is a functional beverage composition comprising a complex of theanine and fermentated extract of rice bran containing GABA. The functional beverage composition of the present invention is effective in improving concentration, learning ability and memory.

Description

FUNCTIONAL BEVERAGE COMPOSITION FOR IMPROVING ABILITY OF LEARNING, CONCENTRATION AND MEMORY

Technical Field The present invention relates to a functional beverage composition for improving concentration, learning ability and memory. More particularly, it relates to a functional beverage composition comprising theanine and fermentated extract of rice bran.

Background Art Theanine was separated in 1950 from gyokuro (a kind of green tea) by Sakato Yajiro. It was named so after Thea sinensisn, the scientific name of the tea and has been identified to be L-glutamate-σ-ethylamide. One of the components enhancing taste of green tea, theanine is an amino acid rich in gyokuro, a high-quality tea. Nitrogen compounds are important components of tea with regard to its taste. While caffeine makes up 50 % of all water-soluble nitrogen compounds, it is theanine, an amino-based nitrogen compound accounting for about 10 %, that contributes to the taste of tea. Theanine is typically comprised in 1-2 % in dry tea leaves, mostly as free amino acid. It takes up 50 % or more of all amino acids present in tea. At most 3 % is present in D-form and most are present in L-form. Theanine is known to significantly increase the level of dopamine when administered orally. Dopamine is an important substance in neurotransmission of brain cells and is known to significantly affect human emotions (Juneja, L.R., Chu, D.D., Okubo, T., and Yokogoshi, H., L-Theanine-unique amino acid of green tea and its relaxation effect in human. Trends in Food Science and Technology, 10,199, 1999). Theanine was listed in the food additives in Japan in July 15, 1963. Currently, it is sold as added in more than 20 kinds of health food products, including beverages, chewing gums, ice cream and jelly, in Japan, Taiwan, etc. GABA, or y-aminobutyric acid (also known as piperidinic acid), is the main ingredient of fermentated extract of rice bran. It has a molecular weight of 103.12 and a melting point of 203 °C and is highly soluble in water. In biosystem, it is produced by an enzyme as carbon dioxide comes off glutamic acid. It had been known to exist in plants from long time ago, but it was in 1950 that GABA was found in brains of animals by an American biochemist Eugene Roberts. Examination of animal tissues revealed that it exists only in the brain of mammals. Since it does suppressive function in the central nervous system of higher animals, it is believed to be a suppressive neurotransmitter. GABA, at once an amino acid and a neurotransmitter, is effective in controlling spasm or depression, reducing blood pressure (thus advantageous in preventing complications caused by diabetes, improving liver and kidney capabilities, preventing obesity and improving climacteric disturbance, asthenia, headache, etc.), facilitating brain cell metabolism (thus improving learning ability), improving memory, relieving stress, relaxing premenstrual nervousness of women and enhancing immune system. Usually, human body produces all the GABA requirements. However, excessive intake of food additives, estrogen, free radicals, salicylates and food additives, low-protein diets and dearth of zinc and vitamin B6 inhibit production of GABA. The daily requirement of GABA to be taken by food is about 500 mg to 4 g. Since it is non-toxic and helps reducing blood pressure, it is used as new functional food supplement in Europe, North America and Japan. Disclosure of Invention It is an object of the present invention to provide a functional beverage composition for improving concentration, learning ability and memory. The above object and other objects can be attained by the present invention as described hereinbelow. To attain the object, the present invention provides a functional beverage composition comprising theanine and fermentated extract of rice bran. The fermentated extract of rice bran may comprise 8-12 % of GABA. Mixing proportion of the theanine and the fermentated extract of rice bran may be 5-20 to 1. The functional beverage composition may comprise theanine having a concentration of 0.5-5 mg/kg and fermentated extract of rice bran. Hereunder is given a detailed description of the present invention. The functional beverage composition of the invention theanine and fermentated extract of rice bran. The fermentated extract of rice bran comprises 8-12 %, preferably 10 %, of GAB A. Mixing proportion of the theanine and the fermentated extract of rice bran is 5- 20 to 1, preferably 20 to 1. The beverage composition comprises a mixture of theanine having a concentration of 0.5-5 mg/kg, preferably 3.5 mg/kg, and fermentated extract of rice bran. The theanine and the fermentated extract of rice bran are prepared as follows. -Preparation of theanine- 1) Enzyme-producing bacterium: Pseudomonas nitroreducens, which produces L-theanine transferase, is cultivated in culture medium to produce the enzyme. 2) Bacterium fixation: Bacterium suspension prepared for use in continuous process is mixed with K-carrageenan to fix the enzyme on a specific support. 3) Enzymatic conversion: The fixed enzyme is filled in a resin column and glutamine and ethylamine solutions are passed to produce L-theanine. 4) Crystallization: The reaction solution is dissolved in hot ethanol (99%) and cooled down for crystallization. Then, it is recrystallized in cold ethanol. 5) Drying: The crystal is dried to obtain theanine. -Preparation of GAB A- 1) Preparation of rice bran extract a) Fermented Korean rice bran (10 % of total amount) is put in an extractor. b) 10 volume equivalents of water is added and extraction is perfoπned by stirring at 60 ^°C for at least 12 hours. c) Insoluble components are removed by centrifuge and the resultant rice bran extract is transferred to a culture apparatus. 2) Cultivation a) The rice bran extract is put in a fermentation system. b) Culture ingredients (sucrose, yeast extract, MSG, etc.) are added. c) Sterilization is performed. d) Lactobacillus sakei is inoculated. e) Cultivation is carried out at 30 °C for at least 48 hours. 3) Sterilization of culture medium After fermentation, the culture medium is sterilized at a temperature of 80 ^°C or above for at least 10 minutes. 4) Removal of solids The culture medium is filtered to remove tyndallized lactobacilli and insoluble solids. 5) Drying Dextrin is added to the culture medium and the culture medium is spray dried to obtain a product having a certain level of GABA. From the resultant comprising theanine and fermentated extract of rice bran, a beverage composition can be prepared according to conventional methods by someone of ordinary skill in the art. Improvement in spatial perception, concentration and memory of animals to which the beverage composition comprising theanine and fermentated extract of rice bran has been administered is measured as follows. 1. Morris water maze test: Improvement in spatial perception and concentration of a mouse to which a functional beverage composition has been repeatedly administered orally for 4 weeks is evaluated by in- vivo effect testing. 2. Passive avoidance test: Improvement in memory of a mouse to which a functional beverage composition has been repeatedly administered orally for 4 weeks is evaluated by in- vivo effect testing. 3. T-maze test: Improvement in memory of a mouse to which a functional beverage composition has been repeatedly administered orally for 4 weeks is evaluated by in- vivo effect testing.

Brief Description of the Drawings FIG. 1 is a graph showing improvement in alpha wave depending on the theanine content. FIG. 2 is a graph showing improvement in alpha wave with time when a beverage composition of the invention comprising 200 mg of theanme and 10 mg of fermentated extract of rice bran was taken. FIG. 3 is a graph showing improvement in spatial perception and concentration of a mouse when a beverage composition of the invention comprising theanine and fermentated extract of rice bran was repeatedly administered orally for 4 weeks (Morris water maze test). FIG. 4 is a graph showing improvement in memory of a mouse when a beverage composition of the invention comprising theanine and fermentated extract of rice bran was repeatedly administered orally for 4 weeks (passive avoidance test). FIG. 5 is a graph showing improvement in memory of a mouse when a beverage composition of the invention comprising theanine and fermentated extract of rice bran was repeatedly administered orally for 4 weeks (T-maze test).

Best Mode for Carrying Out the Invention Hereinafter, the invention is described in further detail through examples. However, the following examples are only for the understanding of the invention and the invention is not limited to or by the examples. [Example 1J -Preparation of theanine- Pseudomonas nitroreducens, which produces L-theanine transferase, was cultivated in culture medium to produce the enzyme. Bacterium suspension prepared for use in continuous process was mixed with K-carrageenan to fix the enzyme on a specific support. The fixed enzyme was filled in a resin column and glutamine and ethylamine solutions were passed to produce L-theanine. The reaction solution was dissolved in hot ethanol (99%) and cooled down for crystallization. Then, it was recrystallized in cold ethanol and dried to obtain theanine. -Preparation of GABA- Fermented Korean rice bran (10 % of total amount) was put in an extractor. 10 volume equivalents of water was added and extraction was perfoπned by stiπing at 60 °C for at least 12 hours. Insoluble components were removed by centrifuge to obtain a rice bran extract. The rice bran extract was put in a fermentation system. Culture ingredients (sucrose, yeast extract, MSG, etc.) were added and sterilization was performed. Lactobacillus sakei was inoculated and cultivation was carried out at 30 ^°C for at least 48 hours. After fermentation, the culture medium was sterilized at a temperature of 80 ^°C or above for at least 10 minutes. The culture medium was filtered to remove tyndallized lactobacilli and insoluble solids. Dextrin was added to the culture medium and the culture medium was spray dried to obtain a product having a certain level of GABA. -Preparation of functional beverage comprising theanine and fermentated extract of rice bran- A beverage composition comprising a complex of the resultant theanine (Taiyo Kagaku Co., Ltd.) and fermentated extract of rice bran (Biovan Co., Ltd.) (mixing proportion = 20 to 1) was prepared with the contents given in Table 1 below. [Table 1)

[Test Example 1] For 10 mg of fermentated extract of rice bran, increase in alpha wave with varying theanine contents (0, 50 and 200 mg/kg) was measured. Optimum proportion of theanine to fermentated extract of rice bran was found to be 5-20 to 1. The most alpha wave was observed when theanine content was 200 mg/kg for 10 mg of fermentated extract of rice bran. [Test Example 2]: Human test For a beverage composition comprising a complex of theanine and fermentated extract of rice bran with a mixing proportion of 20 to 1, random allocation, placebo comparison and double-blind crossover/parallel tests were performed to. identify improvement in relax and mental fatigue relieving (Catholic University of Korea, St. Mary's Hospital, Dream CIS 2001). The result is shown in FIG. 2. 20 healthy adults of ages between 30 and 55 who complaint fatigue lasting at least one month were tested. Double-blind crossover/parallel test was performed while administering testing materials for 7 days. Power proportion of alpha and beta waves was calculated to measure nervousness relieving effect. A test beverage comprising 200 mg of theanine and 10 mg of femientated extract of rice bran (containing 10 % of GABA) was found to increase alpha wave, which is known to be related with mental relaxation and concentration. Particularly, increase in alpha wave was much more prominent when the subject had moderate to severe anxiety. The beverage was also effective in subjects' realization of improvement in mental fatigue. [Test Example 3]: Animal test In-vivo effect of a memory improver was tested by Morris water maze test while repeatedly administering it orally to a mouse for 4 weeks (Chemon Inc., December 2003). The result is shown in FIG. 3. A beverage composition comprising a complex of theanine and of fermentated extract of rice bran with a mixing proportion of 20 to 1 was repeatedly administered orally to a mouse at a dose of 0, 50, 200 and 800 mg/kg for 4 weeks. Morris water maze test was performed for 2 weeks to observe improvement in concentration and memory. General symptoms and animal death were observed. The result is as follows. - General symptoms and animal death No general symptom or animal death caused by administration of test material and control material was observed. - Measurement of duration time Based on the performance of the first day, time duration required to find the platform was observed for 14 days. The groups to which testing materials was administered showed significantly reduced time duration compared with that to which control material was administered. - Measurement of total distance Based on the performance of the first day, distance required to find the platform was observed for 14 days. The test group to which a dose of 200 mg/kg was administered showed significantly reduced distance. The groups to which 50 and 800 mg kg was administered also showed reduced distance. - Measurement of probe trial time duration The time duration at the place where the platform had been was no different for the first probe trial and the second probe trial for the control group. On the other hand, the test groups showed about 20 % increase in the second probe trial. From these result, it can be seen that the groups to which the test material was administered showed improvement in learning ability and memory. [Test Example 4] In-vivo effect of a memory improver was tested by passive avoidance test while repeatedly administering it orally to a mouse for 4 weeks (Chemon Inc., December 2003). The result is shown in FIG. 4. A beverage composition comprising a complex of theanine and of fermentated extract of rice bran with a mixing proportion of 20 to 1 was repeatedly administered orally to a mouse at a dose of 0, 50, 200 and 800 mg/kg for 4 weeks. Passive avoidance test was performed to observe improvement in memory. The result is as follows. - General symptoms and animal death No general symptom or animal death caused by administration of test material and control material was observed. - Measurement of avoidance time The group to which the test material was repeatedly administered at a dose of

50 mg/kg showed about 38 % increased avoidance time compared with the control group, although not statistically significant. The group to which 200 mg/kg was administered showed similar result to that of the control group. The group to which 800 mg/kg was administered showed about 30 % decreased avoidance time. Although the group to which the functional beverage composition was repeatedly administered for 4 weeks at a dose of 50 mg/kg showed possibility of memory improvement, the groups to which 200 and 800 mg/kg showed no improvement. Thus, there was no dose dependency. [Test Example 5] In-vivo effect of a memory improver was tested by T-maze test while repeatedly administering it orally to a mouse for 4 weeks (Chemon Inc., December 2003). The result is shown in FIG. 5. A beverage composition comprising a complex of theanine and of fermentated extract of rice bran with a mixing proportion of 20 to 1 was repeatedly administered orally to a mouse at a dose of 0, 50, 200 and 800 mg/kg for 4 weeks. T-maze test was performed to observe improvement in memory. General symptoms, weight change and T-maze performance were observed. The result is as follows. - General symptoms and animal death No general symptom or animal death caused by administration of test material and control material was observed. - Evaluation of memory Success rate at the final day of administration was 60, 100, 60 and 50 % for the groups to which 0, 50, 200 and 800 mg/kg was administered, respectively. Thus, statistically significant increase was confirmed for the group to which 50 mg/kg was admimstered. From these results, it can be seen that oral administration of the test material to the mouse at a dose of 50 mg/kg is the most efficient in improving memory under the given test conditions. To sum up the animal test results, a mixture of theanine and fermentated extract of rice bran offers the best memory improvement effect at a concentration of 50 mg/kg. For a person weighing 60 kg, 50 mg/kg coπesponds to 3 g/60 kg. Considering that optimum human dose is 1/10 to 1/100 that of mouse due to the difference in intake ratio, 30 mg/60 kg to 300 mg/60 kg is adequate for humans. That is, the optimum dose of theanine and fermentated extract of rice bran for human is 0.5 mg/kg to 5 mg/kg. Thus, adequate dose of theanine and fermentated extract of rice bran for human body is 200 mg and 10 mg, respectively.

Industrial Applicability As apparent from the above description, the composition of the present invention comprising theanine and fermentated extract of rice bran (containing GABA) was confirmed to be effective in improving concentration, learning ability and memory in pre-clinical animal test and human body test. Thus, it is expected to help relieve people from excessive daily-life stresses. While the present invention has been described in detail with reference to the prefeπed embodiments, those skilled in the art will appreciate that various modifications and substitutions can be made thereto without departing from the spirit and scope of the invention as set forth in the appended claims.

Claims

Claims 1. A functional beverage composition comprising theanine and fermentated extract of rice bran.

2. The functional beverage composition of claim 1, the fermentated extract of rice bran comprising 8-12 % of GABA.

3. The functional beverage composition of claim 1, mixing proportion of the theanine and the fermentated extract of rice bran being 5-20 to 1.

4. The functional beverage composition of claim 1, comprising 0.5-5 mg/kg of a mixture of theanine and fermentated extract of rice bran.