Method for manufacturing processed soybean beverage

Abstract

The purpose of the present invention is to provide a novel technology for altering the soybean flavor of a soybean beverage and improving the palatability of the soybean beverage. In the present invention, a nut-like flavor is imparted to a processed soybean beverage obtained according to a method for manufacturing the processed soybean beverage that includes a step for causing laccase to act on the soybean beverage.

Description

Manufacturing method for processed soybean food and drink

[0001] The present invention relates to a method for producing a processed soybean food or drink. More specifically, the present invention relates to a processing technique for imparting a nutty flavor to a soybean food or drink.

[0002] Consumption of soy foods and drinks has increased significantly in recent years due to a variety of factors, including the health boom, addressing allergy issues, religious reasons, and increased demand at home due to the spread of infectious diseases.

[0003] Soy milk, a typical example of soybean food and beverage, includes "soy milk" made only from soybeans with the bean pulp removed and water, with a soybean solid content of 8% by weight or more; "prepared soy milk" with a soybean solid content of 6% by weight or more and containing additives such as sugar and rice oil to make it easier to drink; and "soy milk beverages" with a soybean solid content of 2% by weight or more or 4% by weight and flavored with fruit juice, coffee, etc.

[0004] Soybean foods and beverages such as soy milk have a distinctive grassy smell because they use soybeans as raw ingredients. For example, in the case of foods and beverages such as "prepared soy milk" and "soy milk drinks," which, under the food labeling standards, can contain ingredients other than soybeans and water, the grassy smell is masked using flavorings, sugar, oil, etc. Other methods proposed to reduce the grassy smell include adding palatinose (Patent Document 1) and adding perilla extract (Patent Document 2).

[0005] JP 2003-230365 A JP 2005-253348 A

[0006] Although several methods for masking the unique flavor of soybeans in soybean foods and beverages are known, their effectiveness is still insufficient. On the other hand, as soybean foods and beverages become more popular, it is desirable to further improve the palatability of soybeans in terms of taste while also following the trend toward health.

[0007] Therefore, an object of the present invention is to provide a new technique for changing the soybean flavor of soybean foods and beverages and increasing the palatability of soybean foods and beverages.

[0008] The present inventors have discovered that treating soybean foods and beverages with a multicopper oxidase imparts a nutty flavor and results in a favorable taste. Given that multicopper oxidase is known only to exert a protein cross-linking effect and that no technical relationship is known between the nutty flavor and protein cross-linking, or between soybean foods and beverages and the nutty flavor, the effect of imparting a nutty flavor, which should be unrelated to soybeans, to soybean foods and beverages was extremely unexpected. That is, the present invention provides the following aspects.

[0009] Item 1. A method for producing a processed soybean food or drink, comprising a step of allowing a multicopper oxidase to act on a soybean food or drink. Item 2. The production method according to Item 1, wherein the soybean food or drink is soy milk. Item 3. The production method according to Item 1 or 2, wherein the multicopper oxidase is laccase. Item 4. The production method according to Item 3, wherein the laccase is derived from the genus Trametes. Item 5. The production method according to any one of Items 1 to 4, wherein the soybean food or drink has a protein content of 1 g / 100 ml or more. Item 6. The production method according to any one of Items 1 to 5, wherein the processed soybean food or drink is a beverage with a viscosity of 1 to 20 mPa s. Item 7. A nutty flavor imparting agent for a soybean food or drink, comprising a multicopper oxidase.

[0010] According to the present invention, a nutty flavor can be imparted to a soybean food or drink by treating the soybean food or drink with a multicopper oxidase.

[0011] 1. Method for Producing Processed Soybean Food or Drink The method for producing a processed soybean food or drink of the present invention is characterized by comprising a step of treating a soybean food or drink with a multicopper oxidase. The method for producing a processed soybean food or drink is described in detail below. In this invention, "processing" refers to treatment to impart a nutty flavor, and processed soybean food or drink refers to a soybean food or drink in which a nutty flavor has been imparted to a raw soybean food or drink.

[0012] The soybean food or drink used in the present invention is a beverage (something to drink; soybean drink) or a food (something to eat; soybean food) made from soybeans. The soybean food or drink contains at least crushed cotyledons of soybean seeds. In addition to crushed cotyledons, the soybean food or drink may also contain crushed hypocotyls and / or seed coats.

[0013] The soy beverage may be any liquid composition containing at least crushed cotyledon portions of soybean seeds dispersed in water. In addition to the crushed cotyledon portions, the soy beverage may also contain crushed hypocotyls and / or seed coats dispersed in water. Specific examples of soy beverages include liquids obtained by crushing and dispersing soybeans in water, liquids obtained by removing soybean pulp from liquids obtained by crushing and dispersing soybeans in water (i.e., soy milk), and liquids obtained by redissolving and / or dispersing dried powders (dried soy milk powders) prepared from these liquids in water. Specific examples of soybean foods and beverages include soybean powders, texturized soy protein, and the like.

[0014] These soybean foods and drinks may be used singly or in combination of two or more kinds. Among these soybean foods and drinks, soybean drinks are preferred, and soy milk is more preferred.

[0015] The amount of protein in a soybean food or drink is not particularly limited, but may be, for example, 1 g / 100 ml or more, preferably 2 g / 100 ml or more, more preferably 3 g / 100 ml or more, even more preferably 3.5 g / 100 ml or more, and even more preferably 4 g / 100 ml or more. The upper limit of the range of the protein amount in a soybean food or drink is not particularly limited, but may be, for example, 12 g / 100 ml or less, 10 g / 100 ml or less, 8 g / 100 ml or less, 6 g / 100 ml or less, 5 g / 100 ml or less, or 4.5 g / 100 ml or less.

[0016] The soybean solids content in soybean foods and beverages is not particularly limited, but may be, for example, 2% by weight or more, preferably 4% by weight or more, and more preferably 8% by weight or more. The upper limit of the soybean solids content range in soybean foods and beverages is not particularly limited, but may be, for example, 16% by weight or less, 14% by weight or less, 12% by weight or less, or 10% by weight or less.

[0017] The soybean food or beverage used in the present invention may contain other ingredients in addition to soybeans and water. These other ingredients include quality improvers such as emulsifiers (sucrose fatty acid esters, phospholipids, monoglycerin fatty acid esters, organic acid monoglycerin fatty acid esters, sorbitan fatty acid esters, polyoxyethylene sorbitan fatty acid esters, polyglycerin fatty acid esters, propylene glycol fatty acid esters, etc.), pH adjusters (sodium carbonate, sodium bicarbonate, calcium carbonate, etc.), colorants, flavorings, seasonings (salt, sugar (sucrose), etc.), and secondary ingredients (fruit juice, coffee, etc.). Because the present invention is excellent at imparting a nutty flavor to soybean food or beverages, it is possible to effectively impart a nutty flavor and enhance palatability without the use of additives (e.g., emulsifiers, flavorings, seasonings, secondary ingredients) that have flavor-masking, seasoning, and / or off-taste-masking effects. From this perspective, suitable examples of soybean foods and beverages used in the present invention include soybean foods and beverages that do not contain emulsifiers, flavorings, seasonings, and / or secondary ingredients, and more suitable examples include soybean foods and beverages that do not contain emulsifiers, flavorings, seasonings, and secondary ingredients.

[0018] The multicopper oxidase used in the present invention is a group of enzymes that contain multiple copper atoms in their molecules and oxidize polyphenols, methoxyphenols, diamines, bilirubin, ascorbic acid, and the like with molecular oxygen. The number of copper atoms contained in previously known enzymes is typically 2 to 8, but this number is not particularly limited, as it varies depending on the state of the enzyme preparation at the time of analysis and the analytical method. Examples of enzymes classified as multicopper oxidases include laccase, bilirubin oxidase, ascorbic acid oxidase, and ceruloplasmin.

[0019] These multicopper oxidases may be used alone or in combination of two or more. Among these multicopper oxidases, from the viewpoint of further enhancing the effect of imparting a nutty flavor, laccase and bilirubin oxidase are preferred, and laccase is more preferred.

[0020] Laccase is an enzyme with phenoloxidase activity (EC 1.10.3.2). Specific examples of laccase include laccases derived from microorganisms such as fungi and bacteria, and more specific examples include laccases derived from the genera Aspergillus, Neurospora, Podospora, Botrytis, Collybia, Formes, Lentinus, Pleurotus, Pycnoporus, Pyricularia, Trametes, Rhizoctonia, Rigidoporus, Coprinus, Psatyrella, Myceliophtera, Schtalidium, Polyporus, Phlebia, Coriolus, etc.

[0021] These laccases may be used alone or in combination of two or more. Among these laccases, from the viewpoint of further enhancing the effect of imparting a nutty flavor, preferred are laccases derived from the genus Trametes and laccases derived from the genus Aspergillus (more preferably, laccase derived from Aspergillus oryzae), and even more preferred are laccases derived from the genus Trametes.

[0022] Bilirubin oxidase is an enzyme (EC 1.3.3.5) that acts mainly on bilirubin. Examples of bilirubin oxidase include bilirubin oxidases derived from the genera Penicillium, Myrothecium, Trachyderma, and Bacillus.

[0023] These bilirubin oxidases may be used alone or in combination. Among these bilirubin oxidases, from the viewpoint of further enhancing the effect of imparting a nutty flavor, bilirubin oxidases derived from the genus Myrothecium (preferably Myrothecium verrucaria) and the genus Bacillus (preferably Bacillus subtilis) are preferred.

[0024] The amount of multicopper oxidase used is not particularly limited, and may be, for example, 0.005 mU or more per gram of protein. From the viewpoint of further enhancing the effect of imparting a nutty flavor, the amount of multicopper oxidase used per gram of protein is preferably 0.01 mU or more, more preferably 0.05 mU or more, and even more preferably 0.1 mU or more, 0.5 mU or more, 1 mU or more, 5 mU or more, 10 mU or more, 15 mU or more, 20 mU or more, or 25 mU or more. The upper limit of the range of the amount of multicopper oxidase used is not particularly limited, and may be, for example, 300 U or less, 30 U or less, 3 U or less, 100 mU or less, 40 mU or less, 30 mU or less, 25 mU or less, 20 mU or less, 15 mU or less, 10 mU or less, 5 mU or less, 1 mU or less, or 0.5 mU or less per gram of protein.

[0025] The amount of multicopper oxidase used per gram of soybean solids is, for example, 0.001 mU or more, preferably 0.01 mU or more, more preferably 0.02 mU or more, 0.1 mU or more, 1 mU or more, 5 mU or more, or 10 mU or more. There is no particular upper limit to the range of the amount of multicopper oxidase used, and examples of the amount used per gram of soybean solids include 200 U or less, 20 U or less, 2 U or less, 1 U or less, 200 mU or less, 20 mU or less, 5 mU or less, 1 mU or less, 0.5 mU or less, or 0.05 mU or less.

[0026] Regarding the activity of multicopper oxidase, 20 μL of the enzyme solution was added to 20 μL of a 50 mM solution of the substrate 2,2'-Azino-di-[3-ethylbenzthiazoline sulfonate] (ABTS) and 160 μL of a 125 mM sodium phosphate buffer solution, and the reaction was carried out at 37°C. The absorbance at 405 nm was measured 15 minutes and 0 minutes later. One unit (U) is defined as the amount of enzyme that catalyzes the oxidation of 1 μmol of ABTS per minute.

[0027] In the process of allowing the multicopper oxidase to act on the soybean food or beverage, the multicopper oxidase is added to the soybean food or beverage to prepare a soybean food or beverage composition containing the soybean food or beverage and the multicopper oxidase, and the soybean food or beverage composition is maintained in a heated state to allow a reaction that produces a nutty flavor to proceed.

[0028] The reaction temperature in the step of reacting with the multicopper oxidase can be appropriately determined taking into consideration the optimal temperature of the multicopper oxidase, and is, for example, 10 to 70°C, preferably 35 to 70°C, more preferably 40 to 60°C, and even more preferably 45 to 55°C.

[0029] In the step of reacting the multicopper oxidase, it is also acceptable to use the multicopper oxidase in combination with other enzymes.

[0030] The reaction time in the step of reacting with the multicopper oxidase is not particularly limited and may be, for example, 10 minutes or more, preferably 30 minutes or more, and more preferably 1 hour or more. The upper limit of the reaction time range is not particularly limited and may be, for example, 24 hours or less, 12 hours or less, 8 hours or less, 4 hours or less, or 2 hours or less.

[0031] The step of treating with the multicopper oxidase can be terminated by inactivating the multicopper oxidase used. The method for inactivation is not particularly limited, but examples thereof include inactivation at high temperatures (e.g., 85 to 100°C).

[0032] After the step of reacting with the multicopper oxidase, the soybean food and drink composition is cooled to obtain a processed soybean food and drink. The processed soybean food and drink obtained by the production method of the present invention has a changed soybean flavor and is imparted with a nutty flavor, and can be provided as a highly palatable food and drink. Furthermore, the processed soybean food and drink obtained by the production method of the present invention changes in appearance to a pink color as a result of the reaction that imparts the nutty flavor, and this change in appearance can be particularly noticeable when the processed soybean food and drink is a beverage (processed soybean beverage). In the present invention, a preferred processed soybean food and drink is a beverage (processed soybean beverage), and processed soy milk is particularly preferred.

[0033] Furthermore, when the processed soybean food or drink obtained by the production method of the present invention is a beverage (processed soybean beverage), the processed soybean beverage can be obtained as a low-viscosity liquid despite having been treated with a multicopper oxidase, without the sudden increase in viscosity that would be expected from the protein crosslinking action known for conventional multicopper oxidases, such as gelation. Specific viscosities of processed soybean beverages include, for example, 1 to 20 mPa·s, preferably 1.5 to 12 mPa·s, more preferably 2 to 5 mPa·s, and even more preferably 2.5 to 4 mPa·s. The viscosity is measured at 20°C using a ball-type rotational viscometer with a rotor rotation speed of 1,000 rpm.

[0034] The obtained processed soybean food or drink may be prepared as a dried soybean food or drink through a drying process. When the processed soybean food or drink is a beverage, it can also be prepared as a solid processed soybean composition that, when dissolved and / or dispersed in water, gives a soybean beverage imparted with a nutty flavor. The drying method is not particularly limited, but examples include freeze-drying, vacuum drying, spray drying, etc. Furthermore, the form of the solid processed soybean composition may include powder, fine grains, granules, etc.

[0035] 2. Agent for Imparting Nutty Flavor to Soybean Foods and Drinks As described above, multicopper oxidase can alter the soybean flavor of soybean foods and drinks and impart a nutty flavor to them. Accordingly, the present invention also provides an agent for imparting a nutty flavor to soybean foods and drinks, comprising a multicopper oxidase.

[0036] The types and amounts of materials and components used in the nut-like flavor imparting agent are as shown in the section "1. Method for producing processed soybean food and drink" above.

[0037] The present invention will be specifically described below with reference to examples, but the present invention should not be construed as being limited to the following examples.

[0038] (1) Ingredients Soy milk: unsweetened soy milk, protein concentration 4.15 g / 100 mL, soybean solids content 8% by weight or more Laccase: LC-Y120 (Amano Enzyme Inc.), laccase derived from Trametes genus Laccase: laccase derived from Aspergillus oryzae Bilirubin oxidase: bilirubin oxidase "Amano" 3 (Amano Enzyme Inc.), bilirubin oxidase derived from Myrothecium verrucaria Bilirubin oxidase: bilirubin oxidase derived from Bacillus subtilis Protease: TH-PC10F, protease derived from Geobacillus stearothermophilus Glutaminase: GTSD-100NA, Bacillus amyloliquefaciens-derived glutaminase

[0039] (2) Measurement of Enzyme Activity (2-1) Measurement of Multicopper Oxidase (Laccase, Bilirubin Oxidase) Activity In the following test examples, the enzyme activity of multicopper oxidase was measured by the method described below using 2,2'-Azino-di-[3-ethylbenzthiazoline sulfonate] (ABTS, manufactured by Boehringer Mannheim) as a substrate.

[0040] 20 μL of 50 mM ABTS solution, 160 μL of 125 mM sodium phosphate buffer (pH 7.0), and 20 μL of enzyme solution were added to a microplate and mixed, and the reaction was carried out at 37° C. The absorbance at 405 nm was measured 15 minutes and 0 minutes later. The activity of catalyzing the oxidation of 1 μmol of ABTS per minute was defined as 1 unit (U), and was calculated using the following formula:

[0041]

[0042] (2-2) Protease Activity Measurement Method: 5 mL of 0.6% (w / v) casein solution (0.05 mol / L sodium hydrogen phosphate, pH 8.0) was heated at 37°C for 10 minutes, after which 1 mL of a sample solution containing protease was added and immediately shaken. This solution was then allowed to stand at 37°C for 10 minutes, followed by the addition of 5 mL of trichloroacetic acid test solution (containing 1.8 (w / v)% trichloroacetic acid, 1.8 (w / v)% sodium acetate, and 0.33 mol / L acetic acid), shaking, and again at 37°C for 30 minutes, followed by filtration. The first 3 mL of filtrate was discarded, and 2 mL of the next filtrate was measured. 5 mL of 0.55 mol / L sodium carbonate test solution and 1 mL of Folin's test solution (1→3) were added, shaken well, and allowed to stand at 37°C for 30 minutes. The absorbance AT of this solution (enzyme reaction solution) at a wavelength of 660 nm was measured, using water as a control.

[0043] Separately, 1 mL of a sample solution containing protease was measured, 5 mL of trichloroacetic acid test solution (containing 1.8 (w / v)% trichloroacetic acid, 1.8 (w / v)% sodium acetate, and 0.33 mol / L acetic acid) was added, and the mixture was shaken. 5 mL of the sample solution was then added, and the mixture was immediately shaken and left to stand at 37°C for 30 minutes. Except for this, the mixture was treated in the same manner as the enzyme reaction solution described above (blank), and the absorbance AB was measured.

[0044] The amount of enzyme that causes an increase in the colored substance of Folin's test solution equivalent to 1 μg of tyrosine per minute was defined as 1 unit (1 U).

[0045] 1 mL, 2 mL, 3 mL, and 4 mL of a 1 mg / mL tyrosine standard stock solution (0.2 mol / L hydrochloric acid) were measured, and 0.2 mol / L hydrochloric acid test solution was added to each to make a 100 mL solution. 2 mL of each solution was measured, and 5 mL of 0.55 mol / L sodium carbonate test solution and 1 mL of Folin test solution (1 → 3) were added. The mixture was immediately shaken and left at 37°C for 30 minutes. For each of these solutions, 2 mL of 0.2 mol / L hydrochloric acid test solution was measured and the resulting solution was used as a control. Absorbances A1, A2, A3, and A4 at a wavelength of 660 nm were measured. A calibration curve was created, plotting absorbances A1, A2, A3, and A4 on the vertical axis and the amount of tyrosine (μg) in 2 mL of each solution on the horizontal axis, and the amount of tyrosine (μg) per 2 mL of each solution was calculated.

[0046]

[0047] (2-3) Measurement of Glutaminase Activity In the following test examples, the enzymatic activity of glutaminase was measured using L-glutamine as a substrate by the method described below.

[0048] One mL of the enzyme solution was weighed into a test tube and placed in a 37°C thermostatic water bath for 5 minutes. One mL of 2% (w / v) L-glutamine solution (0.1 mol / L acetate buffer (pH 6.0)) preheated to 37°C was added, mixed, and left for exactly 10 minutes. After leaving the tube, 1 mL of 5% (v / v) perchloric acid test solution was added, mixed, and immediately placed in ice water. After leaving the tube for at least one minute, 1 mL of sodium hydroxide test solution (0.75 mol / L) was added and mixed to form a reaction solution. The L-glutamic acid content of the reaction solution was quantified using the L-glutamic acid measurement kit "Yamasa" NEO (Yamasa Shoyu Co., Ltd.). Under these conditions, the amount of enzyme required to produce 1 μmol of L-glutamic acid per minute was defined as one unit.

[0049] Test Example 1 An enzyme was dissolved in water to prepare an enzyme solution. The enzyme solution was added to unadjusted soy milk so that the enzymes shown in Tables 1 and 2 reached the final concentrations shown, and the mixture was stirred in a 50°C water bath for 90 minutes to react. The enzyme was then inactivated in a 90°C water bath and cooled to room temperature. The resulting processed soy milk was evaluated as follows. The results are shown in Tables 1 and 2.

[0050] (Effect of imparting nutty flavor) Seven monitors drank the processed soy milk and scored the results, with +1 point if they perceived a nutty flavor and 0 point if they did not, and the total score of the scores of the seven monitors was used to evaluate the effect of imparting nutty flavor. The raw material unadjusted soy milk met the standard for "soy milk" of the Japanese Agricultural Standards, and had a grassy smell specific to soybeans but no nutty flavor.

[0051] (Viscosity) The viscosity of the processed soy milk at 25°C was measured using a ball rotational viscometer (manufactured by Kyoto Electronics Manufacturing Co., Ltd., EMS-1000, 1000 rpm, spherical probe φ2 mm).

[0052] (Change in appearance) A sensory visual evaluation was performed to determine whether the appearance (color) of the processed soy milk had changed compared to the appearance (color) of the raw material unadjusted soy milk, and if so, what kind of color had changed.

[0053]

[0054]

[0055] As is clear from Tables 1 and 2, by treating soy milk with laccase, the grassy smell characteristic of soybeans was eliminated and a nutty flavor, which is completely different from that of soybeans, was imparted, resulting in highly palatable processed soy milk (Examples 1 to 6). Moreover, while no monitors responded that they perceived a nutty flavor in any of Comparative Examples 1 to 7, all seven monitors responded that they perceived a nutty flavor in Examples 1 to 6, so the effect of imparting a nutty flavor observed in Examples 1 to 6 was remarkable. Furthermore, only the processed soy milk to which a nutty flavor was imparted by treating the soy milk with laccase tended to have a pink appearance.

[0056] This effect of imparting a nutty flavor and the accompanying discoloration was not observed when other enzymes were used (Comparative Examples 2 to 7), indicating that this effect is unique to the use of laccase.

[0057] Furthermore, the processed soy milk obtained by treating soy milk with laccase did not show the expected significant increase in viscosity, despite the use of laccase, which has a cross-linking effect, and all of the processed soy milk had low viscosity (Examples 1 to 6, especially Examples 1 to 4 and 6).

[0058] Test Example 2 Except for using laccase derived from Aspergillus oryze, bilirubin oxidase derived from Myrothecium verrucaria, or bilirubin oxidase derived from Bacillus subtilis as the enzyme, enzyme treatment was carried out in the same manner as in Test Example 1. As a result, it was confirmed that all of the soy milks after processing exhibited a pink color, similar to Examples 1 to 6 of Test Example 1, and therefore it was possible to infer that the effect of imparting a nutty flavor similar to Examples 1 to 6 was achieved.

Claims

1. A method for producing a processed soybean food or drink, comprising a step of reacting a soybean food or drink with multicopper oxidase.

2. The method according to claim 1, wherein the soybean food or drink is soy milk.

3. The method according to claim 1 or 2, wherein the multicopper oxidase is laccase.

4. The method according to claim 3, wherein the laccase is derived from the genus Trametes.

5. The method according to any one of claims 1 to 4, wherein the protein content of the soybean food or drink is 1 g / 100 ml or more.

6. The method according to any one of claims 1 to 5, wherein the processed soybean food or drink is a beverage having a viscosity of 1 to 20 mPa·s.

7. A nutty flavor imparting agent for soybean food and beverages, comprising multicopper oxidase.