WO2015022911A1 - Method for manufacturing tea extract - Google Patents

Abstract

[Problem] To provide a method for manufacturing a tea extract which exhibits rich taste, flavor and sweetness by decomposing proteins, which are contained in tea leaves and which could not sufficiently be decomposed by a conventional protease treatment extraction, and thereby forming amino acids. [Solution] A method for manufacturing a tea extract, including a step (A) for subjecting tea leaves to first-stage enzymatic treatment, a step (B) for increasing the pH of the obtained product by at least 0.1 after the completion of the step (A), and a step (C) for subjecting the resulting product to second-stage enzymatic treatment after the step (B).

Description

Method for producing tea extracts

The present invention relates to a method for producing an enzyme-treated tea extract. More specifically, a tea extract that can significantly improve the palatability of tea beverages by reducing the bitter and astringent taste peculiar to tea while enhancing the sweetness and umami of tea by blending with tea beverages. It relates to a manufacturing method.

In recent years, products filled with tea beverages in cans or plastic bottles have been provided. Unsweetened tea beverages have gained high support from consumers' sweetness, and their production has increased greatly from 1990 to 2010, and since then has gained stable support from consumers. Has formed a stable market, accounting for a high level of proportion. As a recent trend, tea beverages with strong umami and richness and reduced astringency are preferred.

It is a common practice to use tea extracts as part of the raw materials for producing these tea beverages and for the purpose of improving the flavor. The tea extract is obtained by extracting only a part having a specific effect from tea, and can be prepared in quality according to the form, flavor, purpose, etc. of the final product. The use of tea extracts is simple and advantageous in tea beverage production because the desired effect can be easily obtained by adding desirable ones according to the purpose of the final beverage in tea beverage production. It is a method that brings about an effect.

Various proposals have been made in the method for producing tea extracts, and in particular, as a method for obtaining an extract with strong umami, kokumi, and sweetness, a method for effectively utilizing a large amount of protein present in tea leaves. Can be considered. Although approximately 25% of protein is contained in tea leaves (5th edition food ingredient table), since proteins in tea leaves are insoluble in water, they are not used at all in normal hot water extraction or the like. Since it is expected that if the protein that remains in tea leaves and is not used is decomposed with protease, amino acids are produced and a strong tea extract is expected, various attempts have been made. I came. For example, Patent Document 1 proposes a method of treating tea leaf extraction residue with cellulase and protease.

However, since the protein of tea leaves is strongly bound to tannin, release of so many amino acids was not observed even when protease was allowed to act alone. Therefore, the present applicant previously extracted tea leaves in the presence of protease and tannase, thereby decomposing tannin, which is an inhibitory factor of the action of protease, and making protease easy to act on protein. An invention has been proposed and disclosed in which a tea extract having a strong umami and rich taste and a little astringency can be obtained (Patent Document 2). However, even with this method, many of the proteins in the tea leaves still remain undegraded, and it cannot be said that the proteins in the tea leaves are sufficiently utilized.

As another proposal, green tea leaves are extracted with water in the presence of protease, and the resulting extract is further treated with protease (Patent Document 3). Catechin is once extracted and removed by high-temperature extraction. Protease is allowed to act on the tea leaf residue and extracted, and the total amount of amino acids is 2.5% by mass or more based on the tea leaf-derived solid content in the tea extract by combining the first extract and the subsequent extract. A method for obtaining a tea extract, wherein the ratio of the total amount of catechins is 15.0% by mass or less (Patent Document 4), containing at least one of the group consisting of cellulase and hemicellulase, pectinase, and tannase A method for producing a tea leaf extract (Patent Document 5) is proposed in which an enzyme group further containing protease and a tea leaf are mixed with the enzyme group to be processed, and the tea leaf is subjected to enzymatic decomposition extraction treatment. It has made the results of it, but did not say from what has been fully utilize effectively the protein still in the tea leaves.

Japanese Patent Laid-Open No. 4-228028 JP 2003-144049 A JP 2008-67631 A JP 2009-95333 A JP 2011-50271 A

An object of the present invention is to produce a umami taste, a kokumi taste, a sweet taste by degrading a protein that cannot be fully utilized in a conventional method for producing a tea enzyme-treated extract and remains in a tea leaf extract residue into an amino acid more efficiently than in the past. The purpose is to produce a strong tea extract. In addition, by adding the extract obtained by the method of the present invention to tea beverages, the flavor of tea beverages, such as sweetness and umami, is enhanced, bitterness and astringency are suppressed, and the taste of tea beverages is remarkably increased. The excellent effect that it can be improved is brought about.

When tea leaves are crushed and dispersed in water, the pH is usually in the range of pH 5-6. When this system is treated with an enzyme such as a protease or a saccharide-degrading enzyme, the pH usually decreases. In addition, when the tannase treatment is performed as the enzyme treatment, the pH becomes more acidic due to the generation of gallic acid due to the decomposition of tea tannin (particularly the gallate ester of catechins), and becomes about pH 4-5. In addition, when protease treatment is performed, the pH decreases and often decreases to about 4.3 to 4.8. Furthermore, in the conventional enzyme treatment, a method of adding vitamin C or sodium ascorbate is performed to prevent deterioration, but the enzyme treatment is performed by positively adjusting the pH to the optimum pH of the enzyme. The description is not found. This is presumably because in conventional enzyme-treated extraction of teas, extraction is performed effectively while utilizing the natural flavor of tea, and therefore pH adjustment is not performed.

However, when protease treatment is performed without adjusting the pH in this way, an acidic protease must be selected as the protease, and even if the protease is allowed to act, only the protein that dissolves in the acidic region is degraded by the enzymatic reaction. However, a protein that dissolves in a weakly acidic to weakly alkaline region hardly undergoes an enzymatic reaction and remains in the tea leaf extraction residue without being decomposed.

Therefore, as a result of intensive studies to solve the above problems, the present inventors surprisingly increased the pH of tea leaves treated with protease and tannase after treatment, from weakly acidic to weakly alkaline regions. After the preparation, the protein was treated again with protease, and it was possible to decompose proteins that were dissolved in weakly alkaline to weakly acidic regions, which had not been able to be decomposed until now, and more amino acids were released, resulting in a strong taste. It was found that a green tea extract can be obtained. And when the obtained tea extract was blended with tea beverage, the sweetness and umami of tea beverage were enhanced, the bitterness and taste unique to tea were reduced, and the taste of tea beverage was significantly improved. As a result, the present invention has been completed. Thus, the present invention provides a method for producing a tea extract comprising tannase treatment and protease treatment of tea leaves, comprising the following steps A to C.
Step A: A step of treating the tea leaves with the first stage enzyme,
Process B: After the completion of the process A, a process of increasing the pH by 0.1 or more with respect to the pH at which the process A was performed, and a process C: a process of performing the second stage enzyme treatment after the process B.

According to the present invention, any enzyme that can be used for enzyme treatment of tea leaves in the technical field is not limited to tannase and protease, and the production method for efficiently obtaining tea extracts by the above steps A to C. Can provide
(Step A) Step of enzyme treatment of tea leaves in the first stage,
(Step B) Step of increasing pH by 0.1 or more after completion of Step A,
(Step C) Step of performing the second stage enzyme treatment after Step B,
Can also be provided.

In this case, the suitable pH for the first stage enzyme treatment in Step A is in the range of 4.0 to 6.0, and the suitable pH for the second stage enzyme treatment in Step C is: Assuming that the pH is raised by 0.1 or more with respect to the pH at which the step A is carried out, it can be in the range of 4.2 to 11.0. In this case, a pH adjusting agent can be used for maintaining the pH.

As the enzyme to be used, the enzyme in Step A, which is the first stage enzyme treatment, can contain tannase, and can further contain protease. The enzyme treatment in the second stage of the process C may be used as it is without deactivating the enzyme added in the process A, or an enzyme may be newly added in the process C. At this time, the enzyme to be added may be an enzyme different from the enzyme used in step A. In addition, the enzyme in the process C which is an enzyme treatment of the 2nd step can include a protease. The enzyme in the first stage enzyme treatment of step A and / or the second stage enzyme treatment of step C may include glutaminase and / or asparaginase. Furthermore, in both the first stage enzyme treatment of step A and the second stage enzyme treatment of step C, the enzyme can include a saccharide-degrading enzyme. The tea leaves used in the present invention can be non-fermented tea, semi-fermented tea or fermented tea. Furthermore, the present invention includes a step of protease treatment while maintaining the pH within the range of 4.8 to 11.0 by adding the pH adjuster by completely omitting the first stage enzyme treatment. A method for producing the tea extract is also included.

According to the present invention, compared to conventional tea enzyme-treated extracts, tea leaf proteins that have not been sufficiently used until now are further degraded, the amount of free amino acids is significantly increased, and umami, kokumi, and sweet sweet tea Extract can be obtained. Also, by blending the extract with tea beverages, the umami, kokumi and sweetness of tea beverages can be greatly enhanced, and at the same time it has the effect of reducing bitterness and astringency. The taste of beverages can be significantly improved.

FIG. 1 is a graph showing the transition of the amount of amino acid produced in the product 2 of the present invention (Example 2).

Hereinafter, the present invention will be described in more detail.
Tea leaves that can be used as a raw material in the method of the present invention include fresh leaves obtained from buds, leaves, stems, etc. of tea (scientific name: Camellia sinensis (L) O. Kuntze), which is an evergreen tree of the camellia family, tea made tea, For example, any of non-fermented tea, semi-fermented tea, and fermented tea may be used. Examples of the non-fermented tea include sencha, roasted tea, gyokuro, kabusecha, tencha, bancha, tamago green tea, matcha tea, roasted tea, and the like. Semi-fermented teas include baked tea, iron kannon tea, oolong tea, and fermented teas include black tea, Awaban tea, Goishi tea, Puer tea and the like. In addition, tea such as non-fermented tea, semi-fermented tea, and jasmine tea in which fermented tea is added with flowers can also be used. In addition, brown rice tea in which roasted grains are added to tea can also be used. In particular, non-fermented tea and semi-fermented tea, which are generally said to have a high protein and amino acid content, are suitable.

These tea leaves can be mixed or stirred with water by crushing or cutting to an appropriate size before mixing with water. It becomes. The preferable size of pulverization or cutting is about 0.1 mm to the raw material (unground), but it is preferably 0.2 mm to 20 mm in consideration of difficult appearance and mixing / stirring with water. Furthermore, 0.5 mm to 10 mm is preferable. When the pulverized particle size is less than 0.1 mm, the extract is unfavorable because it has a miscellaneous taste and dislike.

The amount of water to be used is not particularly limited as long as the tea leaves are mixed with water and are physically easy to stir, and it cannot be generally specified because it depends on the nature of the tea leaves and the size of the tea leaves. Usually, 2 to 100 parts by mass can be exemplified with respect to 1 part by mass of tea leaves. However, if there is too little water for tea leaves, stirring and enzyme reaction are difficult to perform, and if there is too much water, the concentration of the extract will decrease, so 5 to 50 parts by weight per 1 part by weight of tea leaves Furthermore, 8 to 20 parts by mass is particularly preferable with respect to 1 part by mass of tea leaves. When the amount of water is less than 2 parts by mass with respect to 1 part by mass of tea leaves, stirring cannot be performed, which is inappropriate for enzyme reaction. In addition, when the amount of water used is more than 100 parts by weight per 1 part by weight of tea leaves, the concentration of the extract becomes thin, and a large amount is required when added to beverages, etc. Even in the case of concentration, it is not preferable because a lot of disadvantages such as a large amount of water must be evaporated. Prior to the enzyme treatment, the tea leaf and water mixture is preferably sterilized at about 60 ° C. to about 121 ° C. for about 2 seconds to about 20 minutes, cooled, and then subjected to the enzyme treatment. In order to prevent oxidative degradation of tea leaves, it is preferable to add ascorbic acid or sodium ascorbate to about 10 ppm to 500 ppm with respect to the total amount of the tea leaf and water mixture.

In the present invention, the mixture of tea leaves and water is first subjected to the first stage enzyme treatment as step A. Next, in step B, the pH is raised by 0.1 or more. Further, after step B, the second stage enzyme treatment is performed as step C. By adopting this series of steps, an efficient and effective enzyme treatment can be performed.

(Process A)
Various enzymes can be used as the enzyme used in the first-stage enzyme treatment in Step A, but tannase can be exemplified most preferably. In addition to tannase, a protease may be used in combination. A large amount of protein is present in tea leaves, but even if proteases are simply allowed to act on tea leaves, the release of amino acids is not so much. This is presumably because the protein is tightly bound to tannin. By allowing tannase to act as the first stage of enzyme treatment, the protein and tannin bonds in tea leaves can be cut off, and proteases and other enzymes can easily act.

Tannase is an enzyme that hydrolyzes a depside bond in which gallic acid is ester-bonded to a hydroxyl group in tannin, for example, an enzyme that hydrolyzes epigallocatechin gallate to epigallocatechin and gallic acid. The tannase can be used in the present invention, specifically, for example, Aspergillus (Aspergillus) genus Penicillium (Penicillium) genus, Rhizopus (Rhizopus) genus Rhizomucor (Rhizomucor) genus Lactobacillus (Lactobacillus) genus, Star Staphylococcus (Staphylococcus) genus Streptococcus (Streptococcus) genus, a tannase-producing bacteria belonging to such Ronepinera (Ronepinella) genus, and solid culture or liquid culture in a conventional manner in a medium usually used for culture of these fungi, the resulting culture And those obtained by purifying the product or its treated product by a conventional method. In addition, commercially available tannase such as tannase-KTFH, tannase-KT05, tannase-KT50 (above, manufactured by Kikkoman Biochemifa); tannase (500 U / g, manufactured by Mitsubishi Chemical Foods); Sumiteam (registered trademark) TAN (New Nippon Chemical Industry Co., Ltd.) can also be used. The amount of tannase used varies depending on the titer, etc., and cannot be generally specified, but is usually 0.1 to 50 U / g, preferably about 0.5 to about 20 U / g, based on the mass of tea leaves. can do.

The pH of the tea leaf aqueous suspension is about 5 to 6 as described above, but the optimum pH of tannase is about 5.0 to 5.5. However, when tannase is allowed to act on tea leaves, gallic acid is produced as described above, so that the pH gradually decreases with the progress of the reaction and becomes about 4.0 to 5.0. During this time, it will pass through within the optimum pH range.

In the first stage enzyme treatment, when tannase is allowed to act, it is not necessary to adjust the pH in consideration of the fact that tannase is likely to act slightly on the acidic side, and the pH during the reaction is adjusted. At least about 4.0 to 6.0. However, it goes without saying that the pH may be adjusted as necessary to maintain the pH within the range of 4.0 to 6.0. The reaction temperature and time for the tannase treatment in the first stage are preferably 20 ° C. to 60 ° C., particularly 25 ° C. to 50 ° C. Examples of the reaction time include 5 minutes to 24 hours, preferably 1 hour to 20 hours, and more preferably 4 hours to 18 hours.

In the first-stage enzyme treatment, in addition to tannase, a protease can be further added to act to decompose proteins in tea leaves. As described above, the pH during the first stage enzyme treatment is about 4 to 6, and the optimum pH of tannase is about 5.0 to 5.5. Therefore, it can be said that the protease added at this time is preferably an acidic protease in consideration of the pH range during action. However, there is no particular limitation when considering the second stage enzyme reaction in Step C without inactivating the protease after raising the pH in Step B, and there are at least one commercially available various proteases. Can be used.

Examples of proteases that can be used include Protease A “Amano” SD, Protease M “Amano” SD, Protease P “Amano” 3SD, Umamizyme G, Peptidase R, Neurase® F, Prozyme, Proleza (registered trademark) ) FG-F, Proteax (registered trademark), Protin SD-NY10, Samoaase (registered trademark) PC10F, Papain W-40 (above, manufactured by Amano Enzyme); Sumiteam (registered trademark) AP, LP, MP, FP, LPL (above, Shin Nippon Kagaku Kogyo Co., Ltd.); Denapsin 2P, Denateam (registered trademark) AP, XP-415, purified papain for food (above, produced by Nagase ChemteX); Orientase (registered trademark) AY, 10NL, 90N, 20A, ONS, Tetrase (registered trademark) S, Nucleicin (Registered trademark) (above, manufactured by HIBI); Morsin (registered trademark) F, PD enzyme, IP enzyme, AO-protease (above, manufactured by Kikkoman Biochemifa); Sakanase (produced by Kaken Pharmaceutical); Protease YP-SS, Pantidase (registered trademark) NP-2, P, Aroase (registered trademark) AP-10 (above, Yakult Pharmaceutical Co., Ltd.); Flavorzyme (registered trademark), Protamex, Neutase, Alcalase (above, Novozymes) ; Coclase (registered trademark) SS, P (above, manufactured by Mitsubishi Chemical Foods); VERON (registered trademark) PS, W, COROLASE (registered trademark) PN-L, N, 7089 (above, manufactured by ABEnzymes); Entilon NBS (Manufactured by Nitto Kasei Kogyo); Protex 7L, Protex 14L (above, Nisuko Japan Co., Ltd.); Actinase (registered trade mark) AS (Kaken Pharma Co., Ltd.); Other animal-derived pepsin, trypsin, or the like can also be mentioned. The protease can be further enhanced by using one or a combination of two or more. The amount of protease used cannot be generally specified depending on the titer and the like, but can be exemplified by a range of 0.01 to 100 U / g based on the mass of tea leaves.

As enzyme treatment conditions other than pH, normal enzyme treatment conditions according to the protease used can be employed. The temperature of the enzyme reaction does not necessarily have to be reacted at the optimum temperature of the enzyme, and in order to prevent flavor deterioration, it may be preferable to carry out the reaction at a slightly lower level. Similar to the tannase treatment, 20 ° C. to 60 ° C., particularly 25 ° C. to 50 ° C. is preferable. Examples of the reaction time include 5 minutes to 24 hours, preferably 1 hour to 20 hours, and more preferably 4 hours to 18 hours.

Also, during the enzyme reaction, about 10 ppm to 500 ppm of ascorbic acid or sodium ascorbate may be added to the total amount of the enzyme extract to prevent oxidative degradation of tea leaf components.

(Process B)
In the present invention, after step A, a step of raising pH as step B is performed. By performing the step of increasing the pH, in the second stage enzyme treatment of the next step C, an enzyme having a characteristic different from that of the first stage enzyme is likely to act, and the entire process is efficient. It can effectively break down tea leaf components, especially proteins. Although the value of the pH to be raised is not particularly limited, it can be 0.1 or more, preferably 0.2 or more, more preferably 0.4 or more, with respect to the pH at which Step A is performed. The above is more preferable, 0.8 or more is particularly preferable, and 1.0 or more is most preferable. By setting the value of increase in pH to a certain large value, in the second stage enzyme treatment, there is a tendency that an enzyme having characteristics different from those of the first stage enzyme tends to act.

As described above, the pH in the first stage enzyme treatment is about 4 to 6, but the pH after being raised in Step B is 4.2 to 11.0, preferably 4.4 to 10.0, More preferably, it can be 4.6 to 9.0, and even more preferably 4.8 to 8.0.

In order to increase the pH in step B, a method of adding a pH adjusting agent can be employed. As the pH adjuster, a general alkali metal salt that can be used as a food additive can be used, and examples thereof include sodium hydrogen carbonate, sodium carbonate, sodium hydroxide, potassium hydroxide, and potassium carbonate. The pH adjuster can be added at once after the completion of the first stage of the enzyme treatment, but is added additionally while measuring the change in pH during the second stage of the enzyme treatment, and the pH is adjusted to 4. It is also possible to employ a method of keeping within the range of 2 to 11.0, preferably 4.4 to 10.0, more preferably 4.6 to 9.0, and even more preferably 4.8 to 8.0. it can. The amount of the pH adjusting agent to be used cannot be generally specified depending on the conditions such as the amount of tea leaf and enzyme used, the enzyme used together, etc., but can be exemplified by about 0.01% to 1% by mass with respect to the tea leaf. .

In addition, between the enzyme treatment of the 1st step | paragraph of the process A and the enzyme treatment of the 2nd step | paragraph of the process C, an enzyme deactivation process may be performed and an enzyme deactivation may not be performed. . As a condition for inactivating the enzyme, a heat treatment at about 60 ° C. to about 121 ° C. for about 2 seconds to about 20 minutes can be employed. When enzyme deactivation is not performed, the enzyme used in the first stage enzyme treatment continues to act in the second stage enzyme treatment of Step C. For example, when the enzyme used as the first-stage enzyme is an enzyme preparation containing a protease that acts in a slightly alkaline region, an action at a pH different from that of the first-stage enzyme treatment can be expected.

(Process C)
In the present invention, after the process B, the process of the second stage enzyme treatment is performed as the process C. By this second-stage enzyme treatment, an enzyme that may have different characteristics from the first-stage enzyme acts, and as a whole, the tea leaf components, particularly proteins, can be decomposed efficiently and effectively. As described above, the pH of the second stage enzyme treatment is 4.2 to 11.0, preferably 4.4 to 10.0, more preferably 4.6 to 9.0, and even more preferably 4.8. Although a range of about 9.0 can be adopted, special attention is required when the pH is particularly high, for example, when the pH is 9 or more. If the pH is 9 or more, the tea leaf components can be decomposed efficiently, but on the other hand, the tea leaf extract is browned or a dough-like odor occurs due to decomposition. May become prominent.

As the enzyme in the second stage enzyme treatment, a protease is preferable, and an enzyme that acts in a slightly alkaline region from a neutral region is particularly preferable. Examples of proteases that can be used include commercially available proteases similar to those described above. The amount of protease used in this step cannot be generally specified due to the titer and the like, as in the first stage enzyme treatment. For example, a range of 0.01 to 100 U / g based on the mass of tea leaves is exemplified. can do.

Also, in the second stage enzyme treatment, about 10 ppm to 500 ppm of ascorbic acid or sodium ascorbate may be added to the total amount of the enzyme extract to prevent oxidative degradation during the enzyme reaction.

In addition, the reaction temperature and time can also be employed under normal enzyme treatment conditions depending on the protease used. The temperature of the enzyme reaction does not necessarily have to be reacted at the optimum temperature of the enzyme, and it may be preferable to perform the reaction at a slightly lower temperature in order to prevent flavor deterioration. For example, 20 to 60 ° C. can be exemplified, In particular, 25 ° C to 50 ° C is preferable. Examples of the reaction time include 5 minutes to 24 hours, preferably 1 hour to 20 hours, and more preferably 4 hours to 18 hours.

Further, in the present invention, when protease and tannase are used in step (A), tea extraction with stronger taste is achieved by allowing glutaminase and / or asparaginase to act in step (A) and / or step (C). Products, in particular green tea extract.

Glutaminase is an enzyme that has the activity of hydrolyzing glutamine or theanine into glutamic acid. Specifically, filamentous fungi or E. coli having the ability to produce glutaminase are cultured according to a conventional method, and the resulting culture is purified by a conventional method. Can be mentioned. Further, commercially available glutaminases such as Glutaminase (from Fluka: derived from filamentous fungi), Glutaminase (from SIGMA: derived from E. coli), Glutaminase Daiwa C100S (Daiwa Kasei Co., Ltd .: derived from filamentous fungi), Glutaminase Daiwa C300S (Yamato) Kasei Co., Ltd .: derived from filamentous fungi), Glutaminase Daiwa C100M (manufactured by Daiwa Kasei Co., Ltd .: derived from filamentous fungi), Sumiteam OP (manufactured by Shin Nippon Chemical Co., Ltd .: derived from filamentous fungi), and the like may be used. The amount of glutaminase used varies depending on the titer and the like, and examples thereof include a range of 0.001 to 100 U / g based on the weight of tea raw materials. Some commercially available glutaminases do not act on theanine but act only on glutamine, such as Sumiteam GT (manufactured by Shin Nippon Chemical Co., Ltd .: derived from filamentous fungi).

Asparaginase is an enzyme having the activity of hydrolyzing asparagine into aspartic acid. Specifically, filamentous fungi and Escherichia coli capable of producing asparaginase are cultured according to a conventional method, and the resulting culture is purified by a routine method. Things can be mentioned. Further, commercially available asparaginase, for example, asparaginase (manufactured by DSM Nutrition Japan Co., Ltd .: derived from filamentous fungi) may be used. The amount of asparaginase used varies depending on the titer and the like, and examples thereof include a range of 0.001 to 100 units / g based on the weight of tea raw materials.

The free amino acids in tea leaves, or teas made from tea leaves, especially free amino acids in green tea, usually have a large proportion of theanine as the main component, but glutamic acid and aspartic acid also have a significant proportion, Glutamine and asparagine are not so much contained in normal tea leaves. On the other hand, when tannase and protease are allowed to act in step (A) to decompose the constituent proteins present in tea leaves, theanine is not produced at all, and glutamic acid and aspartic acid are not produced in large amounts, but glutamine and asparagine are produced in large quantities. To generate. Glutamic acid and aspartic acid are considered to be amino acids that greatly contribute to the umami taste of tea. In step (A), tannase and protease are allowed to act on tea leaves to produce glutamine and asparagine, which are converted into steps (A) and / or By causing glutaminase and / or asparaginase to act in step (C), a glutamic acid and / or aspartic acid can be produced to obtain a green tea extract having a strong taste that could not be obtained by a conventional method.

Theanine is actually considered to be a component that does not contribute much to umami, and by converting theanine to glutamic acid, umami can be enhanced, but theanine is a component unique to tea and has various excellent properties. Component with high functionality. Therefore, when it is desired to use theanine effectively, glutaminase that does not act on theanine but acts only on glutamine can be used as glutaminase.

In the present invention, a saccharide-degrading enzyme can be used in combination in either the first stage or the second stage. Extracting teas with a richer sweetness and richer taste by causing sugar-degrading enzymes to act on tea leaves to break down cellulose, hemicellulose, pectin, etc. in tea leaves to produce monosaccharides, disaccharides, oligosaccharides, etc. You can get things.

Examples of the saccharide-degrading enzyme that can be used in the present invention include enzymes that act on polysaccharides such as pectinase, cellulase, hemicellulase, mannanase, xylanase, and amylase to produce monosaccharides and oligosaccharides. However, it is not limited to these.

Pectinase is also called polygalacturonase, pectin enzyme, polymethylgalacturonase, and pectin depolymerase, and is an enzyme that hydrolyzes α-1,4 bonds such as peclinic acid, pectin, and pectic acid. Pectinase is known to be contained in bacteria, molds, yeasts, higher plants, snails, etc., and pectinases collected from organisms including these can be widely used in the present invention. Commercially available pectinase preparations can also be used. For example, pectinase PL “Amano”, pectinase G “Amano” (manufactured by Amano Enzyme); Pectinase-GODO (manufactured by Godo Shusei); sucrase (registered trademark) A, N, S (manufactured by Mitsubishi Chemical Foods) ); Sumiteam (registered trademark) AP-2, SPC, SPG, MC, PX, liquid Sumiteam AP-2 (above, manufactured by Shin Nippon Chemical Industry Co., Ltd.); pectinase XP-534 (manufactured by Nagase ChemteX); pectinex ( (Registered trademark), Pectinex Ultra SP-L, Ultrazyme (registered trademark), Vinozyme, Citorozym (registered trademark), Peelzyme (registered trademark) (above, manufactured by Novo Nordisk Bioindustry); Cellulosin (registered trademark), PE60 , PEL, soluble pectinase T (above, manufactured by HBI) Pectinase SS, pectinase HL (manufactured by Yakult Pharmaceutical Industry Co., Ltd.) and the like can be exemplified.

The amount of pectinase used is usually less than the active unit because a pectinase preparation usually contains multiple types of enzymes, and is usually about 0.01% to about 5% by weight, preferably about A range of 0.1% by mass to about 2% by mass can be exemplified.

Cellulase is an enzyme that hydrolyzes the glycosidic bond of β-1,4-glucan (for example, cellulose). Cellulose is a kind of polysaccharide in which D-glucose is linked without branching by β-1,4 bonds, and the number of glucose is said to be about 5,000. It is a major component of plant cell walls and is highly hydrophilic but insoluble in water. Cellulases include an endoglucanase that cleaves cellulose from the inside of the molecule, and an exoglucanase (cellobiohydrolase) that decomposes from either the reducing end or non-reducing end of a sugar chain to release cellobiose. Also, commercially available cellulases often contain β-glucosidase and release glucose. Cellulase that can be used in the present invention is not particularly limited as long as it has an activity of decomposing cellulose, and any cellulase preparation can be used. Examples of commercially available cellulase preparations include cellulase T “Amano”. ”, Cellulase A“ Amano ”(manufactured by Amano Enzyme); Doricerase (registered trademark) KSM, Multifect (registered trademark) A40, Cellulase GC220 (manufactured by Genencor Kyowa); Cellulase GODO-TCL, Cellulase GODOTCD-H, Cellulase (registered trademark), Cellulase XL-522 (all manufactured by Nagase ChemteX); Cellsoft (registered trademark); Cellulase (manufactured by Toyobo Co., Ltd.); Cellulase (registered trademark); Trademark), Denimax (registered trademark) ( Upper Novozymes); Cellulosin (registered trademark) AC40, Cellulosin (registered trademark) AL, Cellulosin (registered trademark) T2 (above made by HIBI); Cellulase “Onozuka” 3S, Cellulase Y-NC (above made by Yakult Pharmaceutical Co., Ltd.) ); Sumiteam (registered trademark) AC, Sumiteam (registered trademark) C (manufactured by Shin Nippon Kagaku Kogyo Co., Ltd.); Enchiron CM, Enchiron MCH, Biohit (manufactured by Nitto Kasei Kogyo Co., Ltd.) and the like. The amount of cellulase used is usually about 0.01% to about 1% by weight, preferably about 0.1% to about 0.5% by weight, based on the original tea leaves.

Hemicellulase is an enzyme that degrades hemicellulose. Hemicellulose is a polysaccharide other than cellulose and pectin among the polysaccharides that constitute the cell walls of land plant cells. Furthermore, it forms a hydrogen bond with cellulose and a covalent bond with lignin, and serves to reinforce the cell wall. There are many types of hemicellulases that have a structure in which sugars in the side chains are bound to sugars in the main chain that is the skeleton.

Examples of hemicellulase include glucanase, mannanase, α-galactosidase, galactanase, xylanase, arabinase, polygalacturonase, etc., and an enzyme having a plurality of activities for decomposing these various sugar bonds. Can also be taken. Commercially available hemicellulases include, for example, hemicellulase “Amano” (manufactured by Amano Pharmaceutical Co., Ltd.); Bakezyme (registered trademark) HS2000, Bakezyme (registered trademark) IConc (referred to as “Shibel Hegner”, Japan); Cellulosin (registered trademark) HC100, Cellulosin (registered trademark) HC, Cellulosin (registered trademark) TP25, Cellulosin (registered trademark) B, Hemicellulase M (above, manufactured by HTV Corporation); Sumiteam (registered trademark) X (New) Nippon Chemical Industry Co., Ltd.); VERON191, VERON393 (above, manufactured by Lame Enzyme) and the like. The amount of hemicellulase used can be exemplified by the range of about 0.01% to about 1% by weight, preferably about 0.1% to about 0.5% by weight, based on the original tea leaves.

Amylase is an enzyme that converts amylose and amylopectin in starch into glucose, maltose and oligosaccharide by hydrolyzing glycosidic bonds. Amylases include α-amylase, β-amylase, and glucoamylase. α-Amylase is an enzyme that cleaves α-1,4 bonds of starch and glycogen irregularly to produce polysaccharides or oligosaccharides. β-Amylase is an enzyme that breaks down starch and glycogen into maltose. Glucoamylase is an enzyme that produces glucose by decomposing α-1,4 bonds at the non-reducing ends of sugar chains. Of these amylases, glucoamylase is particularly preferred. Glucoamylase is an enzyme that breaks down the α-1,4 bond at the non-reducing end of the sugar chain to produce glucose, so that it has a strong effect on enhancing sweetness because it produces glucose with strong sweetness when it acts on tea leaves. it is conceivable that. Examples of commercially available glucoamylases include Gluc® (registered trademark) SG, Gluczyme (registered trademark) AF6, Gluczyme (registered trademark) NL4.2, Glucamylase for brewing “Amano” SD (above, manufactured by Amano Enzyme); -ANGH (manufactured by Godo Shusei); Cochlase (registered trademark) G2, Cochlase (registered trademark) M (above, manufactured by Mitsubishi Chemical Foods); Optidex L (manufactured by Genencor Kyowa); Sumiteam (registered trademark), Sumiteam ( (Registered trademark) SG (above, manufactured by Shin Nippon Chemical Industry Co., Ltd.); Glucoteam (registered trademark) # 20000 (manufactured by Nagase ChemteX); AMG, Sun Super (above, manufactured by Novozymes Japan); Glutase AN (HTV) Made by the company); UNIASE (registered trademark) K, UNIASE (registered trademark) 2K, UNIASE (registered) 30), Uniase (registered trademark) 6.0F (above, manufactured by Yakult Pharmaceutical Co., Ltd.); Magnax (registered trademark) JW-201 (manufactured by Nitto Kasei Kogyo Co., Ltd.); Grindomil (registered trademark) AG (Danisco Japan) Manufactured). The amount of amylase used can be exemplified by the range of about 0.01% by mass to about 1% by mass, preferably about 0.1% by mass to about 0.5% by mass relative to the original tea leaves. .

Glucanase is an enzyme that hydrolyzes glucan in a broad sense. Glucan is a polymer in which glucose is linked by glycosidic bonds, and there are α-1,4, α-1,6, β-1,3, β-1,4, β-1,6, etc. . There are cases where two binding modes coexist in one glucan, but α type and β type are not mixed. They are called α-glucan and β-glucan, respectively. is there. A typical substance of α-glucan is starch (α-1,4), and a typical substance of β-glucan is cellulose (β-1,4). Glucanase often refers to a substance excluding amylase and cellulase in a narrow sense, and an enzyme that degrades β-glucan (a polymer of glucose by β-1,3, β-1,4, β-1,6 bonds). In other words, the glucanase referred to in the present invention means an enzyme that degrades β-glucan. Commercially available glucanases include, for example, Finizyme (registered trademark), Ultraflo (registered trademark), Viscozyme (registered trademark), Glucanex, Selemix (manufactured by Novozymes Japan); Multifect (registered trademark) BGL Β-glucanase 750L (manufactured by Genencor Kyowa); Tunicase (registered trademark) FN (Daiwa Kasei); glucanase (ICN Biochemical Inc. (California, USA)). The amount of glucanase used can be exemplified by the range of about 0.01% to about 1% by weight, preferably about 0.1% to about 0.5% by weight, based on the original tea leaves.

Mannanase is an enzyme that performs a reaction to hydrolyze the β-1,4-D-mannopyranoside bond. Commercially available enzymes include, for example, mannanase BGM “Amano”, hemicellulase “Amano” 90, cellulase A “Amano” 3, pectinase PL “Amano” (above, manufactured by Amano Enzyme); β-1,4-mannanase (Yakult Pharmaceutical) Sumiteam (registered trademark) ACH, Sumiteam (registered trademark) AC, Sumiteam (registered trademark) X, Sumiteam (registered trademark) SPC (manufactured by Shin Nippon Chemical Co., Ltd.); Cellulosin (registered trademark) GM5 (Hibiai) Suclase C (manufactured by Mitsubishi Chemical Foods Co., Ltd.) and the like can be exemplified. The amount of mannanase used can be exemplified by the range of about 0.01% to about 1% by weight, preferably about 0.1% to about 0.5% by weight, based on the original tea leaves.

Α-Galactosidase is an enzyme that performs a reaction to hydrolyze α-galactoside bonds such as D-galactopyranosyl- (1 → 6) -α-D-glucopyranoside. Examples of commercially available α-galactosidase include Sumiteam (registered trademark) AGS (manufactured by Shin Nippon Chemical Industry Co., Ltd.). The amount of galactosidase used can be exemplified by the range of about 0.01% to about 1% by weight, preferably about 0.1% to about 0.5% by weight, based on the original tea leaves.

In the present invention, by using a combination of two or more of the above-mentioned pectinases, cellulases, hemicellulases, glucoamylases, glucanases, mannanases and α-galactosidases, which are carbohydrate degrading enzymes, the extract can be more effectively used. Sweetness and umami can be enhanced. In the present invention, the combination of α-amylase and / or β-amylase to degrade starch may lead to enhancement of sweetness and umami. α-Amylase and β-Amylase are particularly effective for cereals with a high starch content.

Commercially available α-amylase preparations include Biozyme (registered trademark) F1OSD, Amylase S “Amano” 35G, Biozyme (registered trademark) A, Biozyme (registered trademark) L (manufactured by Amano Enzyme Co., Ltd.); Sumiteam (registered trademark) L (manufactured by Shin Nippon Chemical Industry Co., Ltd.); Christase (registered trademark) L1, Kryster (registered trademark) P8, Kristase (registered trademark) SD80, Kokugen SD-A Kokugen L, Christase (registered trademark) T10S (manufactured by Daiwa Kasei Co., Ltd.); Biotex L # 3000, Biotex TS, Spitase HS, Spitase CP-40FG, Spitase XP-404 (above, manufactured by Nagase ChemteX) ); Grindoor Mill (registered trademark) A (manufactured by Danisco Japan); BAN, FA Gamil (registered trademark), Termamyl (registered trademark), Novamyl (registered trademark), Maltogenase (registered trademark), Lycozyme Supra, Steinzyme (registered trademark), Aquazyme, Thermozyme (registered trademark), Duramil (registered trademark) (and above) Novazymes Japan Co., Ltd.); Fuctamirase (registered trademark) 30, Fuctamirase (registered trademark) 50, Fuctamirase (registered trademark) 10L, Liquefaction enzyme 6T, Liquefaction enzyme, Requifase L45 (above, manufactured by HB eye); VERONAX, VERONGX, VERONM4, VERONELS (above, manufactured by Higuchi Trading Company); UNIASE (registered trademark) BM-8 (manufactured by Yakult Pharmaceutical Co., Ltd.); ratatase, ratatase RCS, SVA, Magnax JW-121, Sumiteam (registered trademark) A- 10. Sumi team (Registered trademark) AS (manufactured by Shin Nippon Chemical Industry Co., Ltd.); Softagen (registered trademark) 3H (manufactured by Taisho Technos Co., Ltd.); , Genencor Kyowa Co., Ltd.); Bakezyme (registered trademark) P500 (Nihon Shibel Hegner). As β-amylase preparation, Optimalto BBA (manufactured by Genencor Kyowa); β-amylase # 1500, β-amylase L, β-amylase # 1500S (above, manufactured by Nagase ChemteX); (Registered trademark) GL (manufactured by HIBI), UNIASE (registered trademark) L (manufactured by Yakult Pharmaceutical Co., Ltd.), and GODO-GBA (manufactured by Godo Sake). Moreover, an amylase complex enzyme preparation containing all of α-amylase activity, β-amylase activity, and glucoamylase activity can also be used. The amount of amylase used can be exemplified by the range of about 0.01% to about 1% by weight, preferably about 0.1% to about 0.5% by weight, based on the original tea leaves.
As the conditions for the enzyme treatment, the conditions for the enzyme treatment in each of the above steps can be used as they are.

Further, in the present invention, the first-stage enzyme treatment can be omitted completely, and by adding a pH adjuster, the protease treatment can be carried out while maintaining the pH within a slightly higher range than when the pH is not adjusted. . Even if tannase treatment is not performed in the first-stage enzyme treatment, by slightly raising the pH, the bond between the protein and tannin becomes loose, and the protein is likely to act on the protein. In addition to the acidic protease conventionally used for enzymatic degradation of tea leaves, neutral proteases, alkaline proteases and the like are likely to act. The pH at this time is not particularly limited as long as it is higher than unadjusted, but the pH is, for example, 4.8 to 11.0, preferably 5.8 to 9.0, more preferably 6.0 to 8. 5, particularly preferably 7.0 to 8.0.

As the pH adjuster in this case, the aforementioned general alkali metal salts that can be used as food additives, for example, sodium hydrogen carbonate, sodium carbonate, sodium hydroxide, potassium hydroxide, potassium carbonate, and the like can be used. Also, as the protease, at least one of the above-mentioned various proteases can be used, and tannase or a saccharide-degrading enzyme can be allowed to act in combination with protease treatment. Also, the reaction temperature and time can be the same as usual enzyme treatment conditions according to the protease used. For example, 20 to 60 ° C. can be exemplified, and 25 to 50 ° C. is particularly preferable. Examples of the reaction time include 5 minutes to 24 hours, preferably 1 hour to 20 hours, and more preferably 4 hours to 18 hours.

The enzyme-treated product after completion of all enzyme reactions is deactivated at 60 ° C. to 121 ° C. for 2 seconds to 20 minutes, cooled, and separated by employing appropriate separation means such as centrifugation and filter paper filtration. A clear tea extract can be obtained.

The obtained tea extract can be used as it is as the tea extract of the present invention, but by further treatment with PVPP (polyvinylpyrrolidone), activated carbon, etc., the tannin remaining in the tea extract or , Caffeine and polyphenol can be removed, and a tea extract having a refreshing sweetness and umami can be obtained. The added amount of PVPP is preferably 5% by mass to 100% by mass, particularly 10% by mass to 50% by mass, based on the solid content of the extract. If it is less than 5% by mass, the effect of improving the taste cannot be expected so much, and if it exceeds 100% by mass, the flavor of the tea itself may be impaired. The treatment with PVPP cannot be generally specified depending on the desired flavor of the tea extract, but for example, a method of stirring for about 10 minutes to about 2 hours in a temperature range of about 10 ° C. to about 50 ° C. is exemplified. be able to. When processing with PVPP or after processing, blending with sodium ascorbate is effective in preventing deterioration of the flavor. The compounding amount of sodium ascorbate is not particularly limited, and examples thereof include about 0.005% by mass to about 0.5% by mass based on the mass of the tea extract.

Thereafter, the obtained tea extract can be made into a concentrated liquid form by employing an appropriate concentration means, for example, vacuum concentration, reverse osmosis membrane concentration, freeze concentration and the like, if desired. The degree of concentration is not particularly limited, but in general, Bx is 3 ° to 80 °, preferably 8 ° to 60 °, more preferably 10 ° to 50 °.

Thus, the tea extract obtained in this way can mix | blend with a tea drink, for example, and can enhance the sweetness and umami taste greatly, and can reduce the bitterness and astringency which tea drink has. The blending ratio cannot be generally specified due to the difference in the required flavor, but is 0.01% by mass to 90% by mass, more preferably 0.1% by mass to 80% by mass.

In addition to tea beverages, milk beverages, functional beverages, sweets such as candy, cookies, cakes, and jelly can also be blended. Incorporating into the milk beverage, functional beverage, confectionery and the like not only imparts a tea flavor, but also enhances the sweetness and umami which they conventionally have.
Hereinafter, preferred embodiments of the present invention will be described in more detail with reference to Examples, Comparative Examples, and Reference Examples, but the present invention is not limited thereto.

In the following examples,% display is described based on mass.

Example 1 (After enzyme treatment without pH adjustment, pH is adjusted to 8.0 and protease treatment)
To an aqueous solution in which 0.15 g of sodium ascorbate was dissolved in 650 g of ion-exchanged water at 75 ° C., 50 g of commercially available No. 1 tea leaves from Shizuoka crushed with a hammer mill (screen 1.2 mm) were added, and sterilized at a temperature of 80 ° C. Immediately cooled to 45 ° C. The pH at this stage was 5.6. To this was added 0.5 g of tannase (Tannase manufactured by Mitsubishi Chemical Foods), and after stirring for 10 minutes, 0.5 g of protease M (protease manufactured by Amano Enzyme) was further added, followed by stirring at 45 ° C. for 8 hours. (Process A). The pH after the reaction was 4.5. After completion of the first stage reaction, the mixture was sterilized by heating at 90 ° C. for 5 minutes, immediately cooled to 45 ° C., and adjusted to pH 8.0 by adding a 10% aqueous sodium hydroxide solution (step B). Further, 0.5 g of Sumiteam LP (protease manufactured by Shin Nippon Chemical Industry Co., Ltd.) was added thereto, stirred for 10 minutes, and allowed to stand at 45 ° C. for 16 hours (Step C). The pH after the reaction was 6.82. After completion of the reaction, solid-liquid separation was performed, and the separated liquid was sterilized by heating at 90 ° C. for 1 minute, cooled, concentrated under reduced pressure to B × 15 ° using a rotary evaporator, cooled to 20 ° C., and then at 800 × g. Centrifugation was carried out for 10 minutes to remove the precipitate, and 158 g of the green tea extract of the present invention (Product 1 of the present invention) was obtained (tea leaf yield 316%, pH 6.75, Bx 15.0 °).

Example 2 (After tannase, protease, pectinase and cellulase treatment without pH adjustment, the pH was adjusted to 8.0 and protease treatment)
To 650 g of 75 ° C. ion-exchanged water, 0.15 g of sodium ascorbate and 50 g of commercially available No. 1 tea leaves crushed with a hammer mill (screen 1.2 mm) were added, sterilized at 80 ° C. and immediately cooled to 45 ° C. . The pH at this stage was 5.6. To this was added 0.5 g of tannase (Mitsubishi Chemical Foods), and after stirring for 10 minutes, 0.5 g of Sumiteam AP2 (Pectinase made by Shin Nippon Chemical Industry Co., Ltd.), 0.5 g of cellulosin AC40 (manufactured by HBI) and Protease M (Protein manufactured by Amano Enzyme) (0.5 g) was added, and the mixture was stirred at 45 ° C. for 8 hours (step A). The pH after the reaction was 4.5. After completion of the first stage reaction, the mixture was sterilized by heating at 90 ° C. for 5 minutes, immediately cooled to 45 ° C., and adjusted to pH 8.0 by adding a 10% aqueous sodium hydroxide solution (step B). Further, 0.5 g of Sumiteam LP (protease manufactured by Shin Nippon Chemical Industry Co., Ltd.) was added thereto, stirred for 10 minutes, and allowed to stand at 45 ° C. for 16 hours (Step C). The pH after the reaction was 6.82. After completion of the reaction, solid-liquid separation was performed, and the separated liquid was sterilized by heating at 90 ° C. for 1 minute, cooled, and then concentrated under reduced pressure to Bx15 ° using a rotary evaporator, cooled to 20 ° C., and 800 × g The mixture was centrifuged for 10 minutes to remove the precipitate, and 197 g of the green tea extract of the present invention (Product 2 of the present invention) was obtained (tea leaf yield 394%, pH 6.61, Bx 15.0 °).

Example 3 (Protease treatment with pH adjusted to 8.0)
To an aqueous solution in which 0.15 g of sodium ascorbate was dissolved in 650 g of ion-exchanged water at 75 ° C., 50 g of commercially available No. 1 tea leaves from Shizuoka crushed with a hammer mill (screen 1.2 mm) were added, and sterilized at a temperature of 80 ° C. Immediately cooled to 45 ° C. The pH at this time was 5.6. After adjusting the pH to 8.0 by adding a 10% aqueous sodium hydroxide solution, 0.5 g of Sumiteam LP (protease manufactured by Shin Nippon Chemical Industry Co., Ltd.) was added and the mixture was stirred for 10 minutes and then allowed to stand at 45 ° C. for 16 hours. It was allowed to react. The pH after the reaction was 7.05. After completion of the reaction, solid-liquid separation was performed, and the separated liquid was sterilized by heating at 90 ° C. for 1 minute, cooled, concentrated under reduced pressure to B × 15 ° using a rotary evaporator, cooled to 20 ° C., and then at 800 × g. The precipitate was removed by centrifugation for 10 minutes to obtain 125 g of the green tea extract of the present invention (Product 3 of the present invention) (with respect to tea leaf yield of 250%, pH 6.98, Bx 15.0 °).

Comparative Example 1 (without enzyme treatment)
To an aqueous solution in which 0.15 g of sodium ascorbate was dissolved in 650 g of ion-exchanged water at 75 ° C., 50 g of commercially available No. 1 tea leaves from Shizuoka crushed with a hammer mill (screen 1.2 mm) were added, and sterilized at a temperature of 80 ° C. Extraction was performed at 45 ° C. for 1 hour. Subsequently, solid-liquid separation is performed, and the separated liquid is sterilized by heating at 90 ° C. for 1 minute, cooled, and then concentrated under reduced pressure to B × 15 ° using a rotary evaporator, cooled to 20 ° C., and then at 800 × g for 10 minutes. Centrifugation was performed to remove precipitates, and 100 g of green tea extract (Comparative product 1) was obtained (200% yield of tea leaves, pH 5.86, Bx 15.0 °).

Comparative Example 2 (Tannase and protease treatment without pH adjustment)
To an aqueous solution in which 0.15 g of sodium ascorbate was dissolved in 650 g of ion-exchanged water at 75 ° C., 50 g of commercially available No. 1 tea leaves from Shizuoka crushed with a hammer mill (screen 1.2 mm) were added, and sterilized at a temperature of 80 ° C. Immediately cooled to 45 ° C. To this, 0.5 g of tannase (Mitsubishi Chemical Foods) was added, and after stirring for 10 minutes, 0.5 g of protease M (protease made by Amano Enzyme) was further added, followed by stirring at 45 ° C. for 8 hours. The pH after the reaction was 4.5. After completion of the reaction, solid-liquid separation was performed, and the separated liquid was sterilized by heating at 90 ° C. for 1 minute, cooled, concentrated under reduced pressure to B × 15 ° using a rotary evaporator, cooled to 20 ° C., and then at 800 × g. Centrifugation was performed for 10 minutes to remove the precipitate, and 117 g of green tea extract (Comparative product 2) was obtained (234% yield of tea leaves, pH 4.51, Bx 15.0 °).

Comparative Example 3 (Protease treatment without pH adjustment after first-stage enzyme treatment)
In Example 1, the same operation as in Example 1 was carried out except that pH adjustment (addition of a 10% aqueous sodium hydroxide solution) was not performed after completion of the first stage reaction, and 140 g of green tea extract (Comparative product 3) (Yield to tea 280%, pH 4.52, Bx 15.0 °).

Example 4 Sensory evaluation (sensory evaluation by adding the product of the present invention and a comparative product to a green tea beverage)
Add 1 kg of green tea leaves from Shizuoka Prefecture to 20 kg of ion-exchanged water heated to 80 ° C., slowly stir for 5 minutes, separate the tea leaves using a 40 mesh wire net, cool the separated liquid to 20 ° C., and extract 14 kg of liquid was obtained, 7.0 g (500 ppm) of sodium ascorbate was added, 2 filtered paper (manufactured by ADVANTEC Co., Ltd .: retention particle size 5μ) to obtain a green tea beverage stock solution (analyzed value of green tea beverage stock solution; Bx: 2.22 °, pH: 6.4, tannin content (iron tartrate method) ): 0.44%, amino acid content: 0.071%). This was subdivided, diluted 10 times (mass ratio) with ion-exchanged water, and prepared by adding 0.5% of each of the inventive products 1 to 3 and comparative products 1 to 3 to the diluted solution. After sterilization by heating for 30 seconds at 50 ° C., the solution was cooled to 88 ° C. and filled into a 500 ml plastic bottle, held for 2 minutes, then cooled to room temperature (25 ° C.) to obtain a green tea beverage containing a plastic bottle.

The above 10 bottled green tea beverages were evaluated by 10 well-trained panelists. Evaluation is very good: 10 points, good: 8 points, slightly good: 6 points, slightly bad: 4 points, bad: 2 points, very bad 0 points for bitter astringency, sweetness, umami, and balance, respectively Was written. Table 1 shows the average points and average contents of comments.

As shown in Table 1, after treatment with tannase and protease at pH 4 to 6 (actual value 5.6) as step (A), pH was adjusted to 8.0 as step (B), and protease treatment as step (C). The green tea beverage to which the product 1 of the present invention was added has a strong umami, sweetness and richness of green tea, a bitter and astringent taste, a good balance of the whole flavor, and a taste like high-quality matcha. There was a very good evaluation. Furthermore, after the treatment with tannase, protease, pectinase and cellulase at pH 4 to 6 (actual value 5.6) as step (A), the pH is adjusted to 8.0 as step (B), and protease treatment as step (C). The green tea beverage to which the present invention product 2 (that is, the product of the present invention 1 and further subjected to a saccharide-degrading enzyme in the first step) has a strong taste and rich taste of green tea and is sweet. However, the bitter and astringent taste is mild and mild, the balance of the whole flavor is good, the taste is like a high-quality matcha tea, and the bitterness, sweetness, umami, and balance of the present invention 1 The evaluation score was higher than that, and it was extremely good.

The green tea beverage to which the present invention product 3 which has been treated with protease by adjusting the pH to 8.0 has the taste, sweetness and body taste of green tea, and has a bitter and astringent taste, but is a good evaluation that it is not so noticeable. The evaluation score was somewhat inferior to the products 1 and 2 of the present invention, but the result was good to some extent.

On the other hand, the green tea beverage to which the comparative product 1 which has not been subjected to the enzyme treatment is added has an evaluation that it has a weak bitter taste and a strong bitter taste, and any of bitterness taste, sweet taste, umami taste and balance. Even the evaluation was low.

The green tea beverage to which the comparative product 2 treated with tannase and protease without adjusting the pH of the tea leaf was added was evaluated that the umami of the green tea was significantly stronger than the green tea beverage to which the comparative product 1 was added. However, the evaluation was slightly lower than the products 1 and 2 of the present invention, and was inferior to the product 3 of the present invention. The bitter and astringent taste was weaker than the green tea beverage to which Comparative Product 1 was added, but it was still quite strong and the sweetness was slightly poor.

In addition, the green tea beverage to which the comparative product 3 was added after the treatment with tannase and protease without pH adjustment and after the enzyme inactivation was added without further pH adjustment and treated with protease, The taste and sweetness were slightly stronger than the added beverage, but the bitter and astringent taste was slightly outstanding and the balance was poor. In the overall evaluation, there was no significant difference compared to the comparative product 2, and it was evaluated compared to the products 1 and 2 of the present invention. Was low.

Example 5 Component Analysis The amino acid compositions of the inventive products 1 to 3 and the comparative products 1 to 3 were analyzed, and solid content yields and amino acids were compared.
Amino acid analyzer: Hitachi High Speed L-8800A
Measurement method: HPLC method by post-column color development using ninhydrin Yield and amino acid analysis value (amino acid concentration) of extracts of the present invention products 1-3 and comparative products 1-3
Is shown in Table 2.
Table 3 shows these values converted to values from tea leaves, and the solid content yield from tea leaves (Bx conversion) and the amount of amino acid extracted from 1 g of tea leaves (mg) are shown in Table 3.

First, comparative product 2 was obtained by performing tannase and protease treatment without adjusting the pH, but about 6 times as many amino acids were extracted as compared to comparative product 1 without any enzyme treatment, and protein in tea leaves. Is decomposed to produce amino acids. On the other hand, the amino acid yield of Comparative Product 3 which was treated with tannase and protease without adjusting the pH, and further treated with protease without adjusting pH after enzyme inactivation was slightly higher than that of Comparative Product 2, It did not increase so much, and it was found that not many amino acids were generated by the second protease treatment.

In contrast, the product 3 of the present invention was treated with protease by adjusting the pH to 8.0, but more amino acids were produced than the comparative product 3 even though tannase treatment was not performed at all. It was. The reason for this is that by making the aqueous dispersion of tea leaves alkaline, the binding between tannin and protein is weakened, and it is presumed that the protease treatment in that state makes it easier for proteases to act on the proteins in tea leaves. . It was also observed that the soluble solids yield from tea leaves increased overall.

Further, the product 1 of the present invention was obtained by treating tannase and protease at pH 4 to 6 as step (A), adjusting pH to 8.0 as step (B), and carrying out protease treatment as step (C) (that is, After the same step as that of Comparative Product 2, the pH was adjusted to 8.0, and the protease treatment was performed), but the amino acid yield was higher than that of Comparative Products 2 and 3, and a large amount of amino acid was obtained by Step (C). Was found to be produced. In addition, the amino acid yield of the product 1 of the present invention is much higher than that of the product 3 of the present invention. By performing the enzyme treatment using tannase and protease in the step (A), the proteolytic degradation in the step (C) is performed. It was recognized that the effect was remarkably enhanced. The reason for this is that in step (A), the tannin in tea leaves is decomposed particularly by the action of tannase treatment, so that the binding between protein and tannin in tea leaves is weakened, and the pH in step (C) is increased. In this protease treatment, it is presumed that the protease easily acts on the tea leaf protein. It was also observed that the yield of soluble solids from tea leaves was further increased overall.

Furthermore, the product 2 of the present invention is a product obtained by allowing a saccharide-degrading enzyme to act in the step (A) of the product 1 of the present invention, but the amino acid yield is further increased compared to the product 1 of the present invention. It was observed that the soluble solids yield from was also further increased overall. It is presumed that cell wall components are decomposed by the action of the saccharide-degrading enzyme, and as a result, the protease is more likely to act.

From the above results, when protease treatment is performed on tea leaves, adding a pH adjuster to increase the pH further degrades the tea leaf proteins that were difficult to be decomposed and significantly increases the amount of free amino acids. Became clear.

Moreover, it was recognized from the result of sensory evaluation in Example 4 that the tea extract having a good flavor has a large amount of amino acid produced, and a tea beverage is obtained by decomposing proteins in tea leaves into amino acids. It was recognized that an extract with high umami, kokumi and sweetness enhancement can be obtained.

Example 6 Transition of Amino Acid Production According to the Present Invention In Example 1 and Comparative Example 3, free amino acids were measured by sampling every 2 hours immediately after the first enzyme addition (0 hours). The measurement method is to sample approximately 1 ml of the reaction solution into a 1.5 ml microtube, and immediately stop the enzyme reaction by boiling the sample solution in a boiling water bath for 5 minutes. After cooling, the sample solution is centrifuged at 15,000 rpm for 5 minutes in a small centrifuge. The supernatant was recovered. The supernatant is appropriately diluted with ion-exchanged water, and 0.6 ml of protein removal solution is added to 0.2 ml of diluted sample solution. After standing for 15 minutes, centrifuge at 15,000 rpm for 5 minutes. Amino acids in the supernatant were quantified by the ninhydrin colorimetric method. The transition of the amount of free amino acid is shown in FIG.

In Example 1, the amount of free amino acids increased rapidly and dramatically due to the step (C) performed 8 hours after the start of the reaction, that is, the enzyme reaction after addition of protease after adjustment to pH 8.0. Admitted. On the other hand, in Comparative Example 3 in which the protease reaction was carried out without adjusting the pH, the free amino acid gradually increased with the passage of time, but after about 16 hours, it was about ½ compared to Example 1. There was a big difference. Therefore, it can be seen that the protein degradation in the tea leaves has progressed dramatically by adjusting the pH to 8.0 after the first-stage reaction and performing the protease treatment.

Examples 7 to 12 (changed pH to be raised in step (B))
To 650 g of 75 ° C. ion-exchanged water, 0.15 g of sodium ascorbate and 50 g of commercially available No. 1 tea leaves crushed with a hammer mill (screen 1.2 mm) were added, sterilized at 80 ° C. and immediately cooled to 45 ° C. . The pH at this stage was 5.6. To this was added 0.5 g of tannase (Mitsubishi Chemical Foods), and after stirring for 10 minutes, 0.5 g of Sumiteam AP2 (pectinase made by Shin Nippon Chemical Industry), 0.5 g of cellulosin AC40 (manufactured by HI) and protease 0.5 g of M (a protease manufactured by Amano Enzyme) was added, and the mixture was stirred at 45 ° C. for 8 hours (step A). The pH after the reaction was 4.5. After completion of the first stage reaction, pH 5.0 (Example 7), 5.5 (Example 8), 6.0 (Example) was added by adding a 10% aqueous sodium hydroxide solution without performing a sterilization step. 9), 6.5 (Example 10), 7.0 (Example 11) or 7.5 (Example 12) (Step B). Further, 0.5 g of Sumiteam LP (protease manufactured by Shin Nippon Chemical Industry Co., Ltd.) was added thereto, stirred for 10 minutes, and allowed to stand at 45 ° C. for 16 hours (Step C). After completion of the reaction, solid-liquid separation was performed, and the separated liquid was sterilized by heating at 90 ° C. for 1 minute, cooled, and then concentrated under reduced pressure to Bx15 ° using a rotary evaporator, cooled to 20 ° C., and 800 × g The mixture was centrifuged for 10 minutes to remove the precipitate, and the green tea extract of the present invention (the present products 7 to 12) was obtained.

The pH adjusted in Step B of Examples 7 to 12, the pH of the products 7 to 12 of the present invention, the product yield (% of tea leaves), the amino acid content (%), the caffeine content (%), and the tannin content (%) Table 4 shows.

Comparative example 4 (thing which does not perform process (B) and (C) of Example 7)
In Example 7, after completion of the first stage enzyme treatment reaction (step A), solid-liquid separation was performed, and the separated liquid was sterilized by heating at 90 ° C. for 1 minute, cooled, and then up to B × 15 ° using a rotary evaporator. After concentration under reduced pressure and cooling to 20 ° C., the precipitate was removed by centrifugation at 800 × g for 10 minutes to obtain a green tea extract (Comparative Product 4).

Comparative Example 5 (in Example 7, step (C) was performed without adjusting pH after step (B))
In Example 7, after completion of the first stage enzyme treatment reaction (Step A), 0.5 g of Sumiteam LP (protease manufactured by Shin Nippon Chemical Industry Co., Ltd.) was further added without stirring the pH, and the mixture was stirred for 10 minutes. The mixture was allowed to stand at 16 ° C. for 16 hours (Step C). After completion of the reaction, solid-liquid separation was performed, and the separated liquid was sterilized by heating at 90 ° C. for 1 minute, cooled, concentrated under reduced pressure to B × 15 ° using a rotary evaporator, cooled to 20 ° C., and then at 800 × g. Centrifugation was performed for 10 minutes to remove the precipitate, and a green tea extract (Comparative product 5) was obtained.
Table 4 shows the pH, product yield (% of tea leaves), amino acid content (%), caffeine content (%), and tannin content (%) of Comparative Products 4 and 5.

As shown in Table 4, the product of the present invention 7 in which the pH was raised (Step B) after completion of the first stage of the enzyme treatment (Step A), and then protease treatment was further added to carry out the enzyme treatment (Step C). In each of -12, the product yield was higher than those of Comparative products 4 and 5 (both of which pH was not adjusted during the enzymatic reaction), and in particular, many amino acids were extracted as components. From Table 4, it can be seen that the pH in Step B is about 6.0 (invention product 9) and the best, but even if it is 5.0 (invention product 7, an increase of 0.3), it is a comparative product. In comparison with 5, it was confirmed that large effects (product yield increasing effect, amino acid yield increasing effect) were obtained. From this result, it is expected that a considerable effect can be obtained even if the pH raised in the step B is small (about 0.1).

Example 7 Sensory evaluation (sensory evaluation by adding the product of the present invention and a comparative product to a green tea beverage)
A green tea beverage stock solution was obtained by the same method as in Example 4 (analyzed value of green tea beverage stock solution; Bx: 2.22 °, pH: 6.4, tannin content (iron tartrate method): 0.44%, amino acid Content: 0.071%). This was subdivided, diluted 10 times (mass ratio) with ion-exchanged water, and prepared by adding 0.5% each of the inventive products 7 to 12 and comparative products 4 and 5 to the diluted solution. After sterilization by heating for 30 seconds at 50 ° C., the solution was cooled to 88 ° C. and filled into a 500 ml plastic bottle, held for 2 minutes, then cooled to room temperature (25 ° C.) to obtain a green tea beverage containing a plastic bottle.

The above 10 bottled green tea beverages were evaluated by 10 well-trained panelists. Evaluation is very good: 10 points, good: 8 points, slightly good: 6 points, slightly bad: 4 points, bad: 2 points, very bad 0 points for bitter astringency, sweetness, umami, and balance, respectively Was written. Table 5 shows the average points and average contents of comments.

As shown in Table 5, after the first stage of the enzyme treatment (Step A) was finished, the pH was raised (Step B), and then protease treatment was added to perform the enzyme treatment (Step C) 7 Each of the green tea beverages to which 12 to 12 were added had stronger umami and rich taste of green tea than the green tea beverages to which comparative products 4 and 5 (both were not adjusted for pH during the enzymatic reaction) The result was that the sweetness was extremely strong, the bitter and astringent taste was mild and mild, the overall flavor was well balanced, and it had a taste like high-quality matcha. Therefore, it is recognized that the taste of the beverage to which the extract of the present invention is added is greatly improved by adding the extract obtained by further performing the enzyme treatment after increasing the pH in Step B. It was. Further, from comparison between the comparative product 5 in which the pH is not adjusted in the process B and the product 7 of the present invention in which the pH is increased by 0.3 in the process B, the increase in pH in the process B is 0.3. It was recognized that a large effect (enhancement effect such as umami) was obtained. From the result of this sensory evaluation, it is expected that a considerable effect can be obtained even if the pH raised in Step B is small (about 0.1).

Example 13
In Example 7, in Step C, 0.5 g of glutaminase GT (not acting on theanine made by Shinnippon Chemical Co., Ltd.) was added to 0.5 g of Summin Team LP (protease made by Shinnippon Chemical Co., Ltd.) and glutamine. The same procedure as in Example 7 was carried out except that 0.5 g of glutaminase and asparaginase were added to obtain 202 g of the green tea extract of the present invention (product 13 of the present invention) (404% yield of tea leaves).
Table 6 shows the amino acid composition of Products 7 and 13 of the present invention.

As shown in Table 6, the product 13 of the present invention has a sharp decrease in asparagine and glutamine compared to the product 7 of the present invention, while the aspartic acid and glutamic acid have increased sharply, and the increase in aspartic acid is almost a decrease in asparagine. The increase in glutamic acid was roughly equivalent to the decrease in glutamine. On the other hand, the content of theanine was almost the same. Therefore, it is presumed that the asparagine of the product 7 of the present invention was converted to aspartic acid by the action of asparaginase, and glutamine was converted to glutamic acid, resulting in the numerical value of the product 13 of the present invention.

Example 14
The inventive products 7 and 13 were each made into 2% aqueous solution and evaluated by 10 panelists who were well trained. As a result, all 10 persons judged that the product 13 of the present invention was stronger than the product 7 of the present invention.

Claims (12)

1. A method for producing tea extracts comprising tannase treatment and protease treatment of tea leaves, comprising the following steps A to C:
   Step A: A step of treating the tea leaves with the first stage enzyme,
   Step B: After step A, the step of raising the pH by 0.1 or more with respect to the pH at which step A was carried out,
   Step C: Step of performing the second stage enzyme treatment after Step B.
2. The first stage enzyme treatment is performed while maintaining a pH range of 4.0 to 6.0, and the second stage enzyme treatment is performed while maintaining a pH range of 4.2 to 11.0. The method described in 1.
3. The method according to claim 1 or 2, wherein the enzyme of step A contains tannase.
4. The method according to claim 1 or 2, wherein the enzyme of step A contains a protease.
5. The method according to any one of claims 1 to 4, wherein an enzyme is added in step C.
6. The method according to claim 5, wherein the enzyme added in step C is an enzyme different from that in step A.
7. The method according to any one of claims 1 to 6, wherein the enzyme of Step C contains a protease.
8. The method according to any one of claims 1 to 7, wherein the enzyme of step A and / or step C comprises glutaminase and / or asparaginase.
9. The method according to any one of claims 1 to 8, wherein the enzyme of step A and / or step C comprises a saccharide-degrading enzyme.
10. The method according to any one of claims 1 to 9, wherein the tea leaves are one or more selected from non-fermented tea, semi-fermented tea or fermented tea.
11. A method for producing a tea extract comprising a step of treating a tea leaf with a protease while maintaining the pH within a range of 4.8 to 11.0 by adding a pH adjusting agent.
12. The following steps A to C,
    Step A: A step of treating the tea leaves with the first stage enzyme,
    Step B: Step of increasing pH by 0.1 or more after completion of Step A,
    Step C: Step of performing the second stage enzyme treatment after Step B,
    Of tea extract containing

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