Fermented beer-taste beverage

Abstract

To provide a fermented beer-taste beverage excellent in flavor durability while having a low fermentation degree.SOLUTION: The fermented beer-taste beverage has an appearance fermentation degree of ≤ 50% and an increment of trans-2-nonenal concentration of ≤ 0.1 μ g / L when stored at 20 °C for one month.SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to a fermented beer-taste beverage. [Background technology]

[0002] Known methods for producing low-alcohol fermented beer-flavored beverages include a method in which a fermented beer-flavored beverage is produced in the usual manner and then distilled to reduce the alcohol content, a method in which a fermented beer-flavored beverage with a low degree of attenuation is produced, and a combination of these methods. For example, Patent Document 1 discloses a method for preparing beer with an ethanol content of 0 to 1.0% by volume, which includes preparing a medium containing limited-fermentation beer and having an ethanol content of 0 to 20% by volume, and subjecting the medium to a distillation step, thereby reducing the amount of one or more aldehydes selected from the group consisting of 2-methylpropanal, 2-methylbutanal, 3-methylbutanal, 3-methylthiopropionaldehyde, phenylacetaldehyde, hexanal, trans-2-nonenal, benzaldehyde, and furfural in the medium, thereby reducing the ethanol content, if present, to a range of 0 to 1.0% by volume. [Prior art documents] [Patent documents]

[0003] [Patent Document 1] Special Publication No. 2022-510753 Summary of the Invention [Problem to be solved by the invention]

[0004] It is known that fermented beer-flavored beverages with low attenuation levels have poor flavor durability and are prone to aging during storage. The present inventors have newly discovered that low-attenuation beer-flavored beverages have significantly weaker antioxidant potential. This poor antioxidant potential makes it easier for substances such as trans-2-nonenal, which are responsible for aging odors, to be produced, which is thought to be one of the reasons for the low flavor durability.

[0005] Based on this novel finding, the present invention aims to provide a fermented beer-flavored beverage that has a low degree of fermentation but excellent flavor durability. [Means for solving the problem]

[0006] The present invention includes, for example, the following inventions. [1] The apparent degree of fermentation is 50% or less, A fermented beer-flavored beverage in which the increase in trans-2-nonenal concentration when stored at 20°C for one month is 0.1 μg / L or less. [2] The fermented beer-taste beverage according to [1], wherein the increase is 0.05 μg / L or less. [3] The apparent degree of fermentation is 50% or less, A fermented beer-flavored beverage with a trans-2-nonenal concentration of 0.2 μg / L or less when stored at 20°C for one month or more. [4] The apparent degree of fermentation is 50% or less, A fermented beer-flavored beverage with an ORAC of 5 μmol TE / g or less. [5] The fermented beer-flavored beverage according to any one of [1] to [4], which has an ORAC of 5 μmol TE / g or less. [6] The fermented beer-flavored beverage according to any one of [1] to [5], wherein the sulfite concentration is 5 mg / L or less. [7] The fermented beer-flavored beverage according to any one of [1] to [6], which has an alcohol content of less than 1 v / v%. [8] The fermented beer-flavored beverage according to any one of [1] to [7], having an apparent degree of fermentation of 40% or less. [9] The fermented beer-taste beverage according to any one of [1] to [8], wherein the malt usage ratio is 30% by mass or more and less than 100% by mass.

[10] It comprises at least a preparation step and a fermentation step, A method for producing a fermented beer-flavored beverage includes adjusting the apparent fermentation degree to 50% or less and suppressing the production of precursor compounds that are converted into substances that cause aged odor.

[11] The manufacturing method described in

[10] , wherein suppressing the production of precursor compounds that are converted into substances that cause aging odor is by inhibiting or blocking the activity of LOX-1 in the raw material, or by using a raw material that does not have LOX-1 activity. [Effects of the Invention]

[0007] According to the present invention, a fermented beer-flavored beverage can be provided that has a low degree of fermentation but excellent flavor durability. DETAILED DESCRIPTION OF THE INVENTION

[0008] Hereinafter, embodiments of the present invention will be described in detail, but the present invention is not limited to the following embodiments.

[0009] The fermented beer-taste beverage according to this embodiment has excellent flavor durability despite its low degree of fermentation. Excellent flavor durability may mean, for example, that deterioration due to storage is suppressed, or that an increase in the degree of aging due to storage is suppressed.

[0010] As described in the Examples, the present inventors have newly discovered that beer-taste beverages with a low degree of fermentation have significantly weaker antioxidant power. Weak antioxidant power facilitates the production of trans-2-nonenal, a substance that causes aging odors (e.g., cardboard odor), and this is thought to be one of the reasons for the low flavor durability. The fermented beer-taste beverage of this embodiment achieves the effect of excellent flavor durability despite weak antioxidant power by suppressing the production of precursor compounds that are converted into substances that cause aging odors during the production process of the fermented beer-taste beverage, thereby suppressing the production of substances that cause aging odors.

[0011] One way to suppress the formation of precursor compounds is to inhibit the activity of barley lipoxygenase-1 (LOX-1) in raw materials. LOX-1, a lipid-oxidizing enzyme found in barley, oxidizes linoleic acid to produce 9-hydroperoxyoctadecadienoic acid. 9-Hydroperoxyoctadecadienoic acid is one of the precursor compounds of trans-2-nonenal and is known to be converted to trans-2-nonenal through cleavage by fatty acid hydroperoxide lyase and subsequent isomerization. If the formation of 9-hydroperoxyoctadecadienoic acid can be suppressed by inhibiting LOX-1 activity, it is thought that the formation of trans-2-nonenal can be suppressed even with weak antioxidant activity.

[0012] In this specification, "beer-taste beverage" refers to a beverage that has a beer-like flavor. Examples of beer-taste beverages include, but are not limited to, those classified as beer, happoshu, and other sparkling alcoholic beverages as defined in Article 3 of the Liquor Tax Act (Act No. 6 of 1953). Beer-taste beverages also include beverages and soft drinks (e.g., non-alcoholic beer-taste beverages) that do not fall under the category of sparkling alcoholic beverages under the Liquor Tax Act. The beer-taste beverage according to this embodiment is not limited to the above examples.

[0013] In this specification, a "fermented beer-taste beverage" refers to a beer-taste beverage produced through fermentation using yeast or the like.

[0014] The fermented beer-taste beverage according to this embodiment has an apparent degree of fermentation of 50% or less. The apparent degree of fermentation is a value calculated according to the method described in "8.5 Extract-Related Calculation Method" in the Revised BCOJ Beer Analysis Methods (published by the Brewery Society of Japan, edited by the International Technical Committee of the Brewers Association of Japan (Analysis Committee) and expanded and revised in 2013). Specifically, the value is calculated using the following formula 1: Formula 1: Apparent fermentation degree (%) = {(original extract (mass%) - apparent extract (mass%)) / original extract (mass%)} × 100 Original extract and apparent extract can be measured using the methods described in "7.2 Extract" and "8.2 Apparent (pseudo) extract" of the Revised BCOJ Beer Analysis Methods (published by the Brewery Society of Japan, edited by the International Technical Committee of the Brewers Association of Japan (Analysis Committee), revised and expanded in 2013).

[0015] The apparent fermentation degree of the fermented beer-taste beverage according to this embodiment may be, for example, 49% or less, 48% or less, 47% or less, 46% or less, 45% or less, 44% or less, 43% or less, 42% or less, 41% or less, 40% or less, 39% or less, 38% or less, 37% or less, 36% or less, 35% or less, 34% or less, 33% or less, 32% or less, or 31% or less. There is no particular lower limit for the apparent fermentation degree of the fermented beer-taste beverage according to this embodiment, but it may be, for example, 10% or more.

[0016] The fermented beer-taste beverage according to this embodiment may exhibit an increase in trans-2-nonenal concentration of 0.1 μg / L or less when stored at 20°C for one month. The "increase in trans-2-nonenal concentration when stored at 20°C for one month" refers to the trans-2-nonenal concentration of the fermented beer-taste beverage after one month of storage at 20°C minus the trans-2-nonenal concentration of the fermented beer-taste beverage before storage began. The fermented beer-taste beverage according to this embodiment may exhibit an increase in trans-2-nonenal concentration of 0.09 μg / L or less, 0.08 μg / L or less, 0.07 μg / L or less, 0.06 μg / L or less, or 0.05 μg / L or less when stored at 20°C for one month.

[0017] The trans-2-nonenal concentration in fermented beer-flavored beverages can be measured by solid-phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS) using the standard addition method. More specifically, the analysis can be performed based on the analytical method described in a non-patent document (Vesely et al., J. Agric. Food. Chem., 2003, Vol. 51, pp. 6941-6944).

[0018] The fermented beer-taste beverage according to this embodiment may have an increase in trans-2-nonenal concentration of 0.2 μg / L or less when stored for one month at 30° C. The "increase in trans-2-nonenal concentration when stored for one month at 30° C" refers to the trans-2-nonenal concentration of the fermented beer-taste beverage after one month of storage at 30° C. minus the trans-2-nonenal concentration of the fermented beer-taste beverage before storage began. The fermented beer-taste beverage of this embodiment may have an increment in trans-2-nonenal concentration when stored at 30°C for one month of 0.19 μg / L or less, 0.18 μg / L or less, 0.17 μg / L or less, 0.16 μg / L or less, 0.15 μg / L or less, 0.14 μg / L or less, 0.13 μg / L or less, 0.12 μg / L or less, 0.11 μg / L or less, or 0.1 μg / L or less.

[0019] The fermented beer-flavored beverage of this embodiment may have a trans-2-nonenal concentration of 0.25 μg / L or less, 0.24 μg / L or less, 0.23 μg / L or less, 0.22 μg / L or less, 0.21 μg / L or less, or 0.2 μg / L or less when stored at 20°C for one month or more.

[0020] The fermented beer-flavored beverage according to this embodiment may have a trans-2-nonenal concentration of 0.35 μg / L or less, 0.34 μg / L or less, 0.33 μg / L or less, 0.32 μg / L or less, 0.31 μg / L or less, 0.3 μg / L or less, 0.29 μg / L or less, 0.28 μg / L or less, 0.27 μg / L or less, 0.26 μg / L or less, 0.25 μg / L or less, 0.24 μg / L or less, 0.23 μg / L or less, 0.22 μg / L or less, 0.21 μg / L or less, or 0.2 μg / L or less when stored at 30°C for one month or more.

[0021] The fermented beer-flavored beverage according to this embodiment may have an ORAC of 5 μmol TE / g or less. In this specification, "ORAC" refers to a value indicating antioxidant activity, as assessed by the ORAC (Oxygen Radical Absorbance Capacity Assay) method. Specifically, a radical donor (2,2'-azobis(2-amidinopropane)dihydroxychloride), a fluorescent substance (fluorescein), and a sample (fermented beer-flavored beverage) are mixed, and fluorescence intensity is measured over time. The area enclosed by the curve of fluorescence intensity versus reaction time with the addition of the sample and the curve of fluorescence intensity versus reaction time without the addition of the sample is calculated as antioxidant activity. Antioxidant activity is calculated in the same manner using a standard substance (Trolox) instead of the sample. The antioxidant activity of 1 μmol of the standard substance (Trolox) is defined as 1 μmol TE, and ORAC is the antioxidant activity of the sample expressed as a relative value per unit weight of the sample.

[0022] The ORAC of the fermented beer-taste beverage according to this embodiment may be, for example, 4.5 μmol TE / g or less, 4 μmol TE / g or less, 3.5 μmol TE / g or less, 3 μmol TE / g or less, 2.5 μmol TE / g or less, 2 μmol TE / g or less, 1.5 μmol TE / g or less, 1 μmol TE / g or less, 0.5 μmol TE / g or less, or 0 μmol TE / g. The fermented beer-taste beverage according to this embodiment has excellent flavor durability despite its weak antioxidant power.

[0023] The fermented beer-taste beverage according to this embodiment may have a sulfite concentration of 5 mg / L or less. The sulfite concentration of the fermented beer-taste beverage according to this embodiment may be, for example, 4.5 mg / L or less, 4 mg / L or less, 3.5 mg / L or less, 3 mg / L or less, 2.5 mg / L or less, 2 mg / L or less, 1.5 mg / L or less, 1 mg / L or less, 0.5 mg / L or less, or 0 mg / L. Sulfite is an antioxidant. The fermented beer-taste beverage according to this embodiment has excellent flavor durability even with such a low sulfite concentration.

[0024] The sulfite concentration of fermented beer-flavored beverages can be measured using the method described in "8.13.1 Titration Method" in the Revised BCOJ Beer Analysis Methods (published by the Brewery Society of Japan, edited by the International Technical Committee of the Brewers Association of Japan (Analysis Committee), revised and expanded in 2013).

[0025] The fermented beer-flavored beverage according to this embodiment may or may not contain a barley raw material as a raw material. In this specification, the barley raw material refers to barley or a processed barley product. Examples of barley include barley, wheat, rye, oats, oats, pearl barley, and oats. Examples of processed barley products include barley extract, malt, and malt extract. Barley extract is obtained by extracting barley extract components containing sugars and nitrogen from barley. Malt is obtained by germinating barley. Malt extract is obtained by extracting extract components containing sugars and nitrogen from malt.

[0026] The fermented beer-taste beverage according to this embodiment may have a malt ratio (the proportion of malt in ingredients other than water and hops) of 0% to 100% by mass. The malt ratio may be 10% by mass or more, 20% by mass or more, 30% by mass or more, 40% by mass or more, 50% by mass or more, 60% by mass or more, 65% by mass or more, 66% by mass or more, 67% by mass or more, 70% by mass or more, 80% by mass or more, 90% by mass or more, 95% by mass or more, 99% by mass or more, or 100% by mass. The malt ratio may also be less than 100% by mass, 95% by mass or less, 90% by mass or less, 80% by mass or less, 70% by mass or less, 60% by mass or less, or 50% by mass or less. The fermented beer-flavored beverage of this embodiment tends to have better flavor durability the lower the malt usage ratio, so the malt usage ratio may be, for example, 30% by mass or more but less than 100% by mass, preferably 30% by mass or more but less than 80% by mass, more preferably 40% by mass or more but less than 70% by mass, and even more preferably 50% by mass or more but less than 60% by mass.

[0027] The fermented beer-taste beverage according to this embodiment may or may not contain ingredients other than barley. Examples of ingredients other than barley include grains such as corn, rice, and sorghum; potatoes such as potatoes and sweet potatoes; beans such as soybeans and peas; herbs and spices; and carbohydrate ingredients (sugars) such as starch, grits, and liquid sugar.

[0028] The fermented beer-flavored beverage according to this embodiment may or may not contain hops as an ingredient. In this specification, hops includes, for example, fresh hops, dried hops, hop pellets, and hop extracts, as well as processed hop products such as extracts of low hops, hexahops, tetrahops, and isomerized hops.

[0029] The fermented beer-flavored beverage according to this embodiment may further contain other components (other ingredients) in addition to the above-mentioned components, provided that the effects of the present invention are not impaired. Examples of other ingredients include bittering agents, coloring agents, sweeteners, high-intensity sweeteners, antioxidants, acidulants, flavoring agents, salts, etc. Examples of bittering agents include iso-α acid, caffeine, gentian extract, peptides, theobromine, naringin, bitter persimmon extract, artemisia absinthium extract, and cinchona extract. Examples of coloring agents include caramel color, gardenia color, fruit juice color, vegetable color, and synthetic color. Examples of sweeteners include high-fructose glucose syrup, glucose, galactose, mannose, fructose, lactose, sucrose, glycogen, and starch. Examples of high-intensity sweeteners include neotame, acesulfame K, sucralose, saccharin, saccharin sodium, disodium glycyrrhizinate, cyclamate, dulcin, stevia, glycyrrhizin, thaumatin, monellin, aspartame, and alitame. Examples of antioxidants include vitamin C, vitamin E, and polyphenols. Examples of acidulants include lactic acid, citric acid, succinic acid, acetic acid, tartaric acid, gluconic acid, malic acid, and phosphoric acid. Examples of salts include table salt, potassium acid phosphate, calcium acid phosphate, ammonium phosphate, magnesium sulfate, calcium sulfate, potassium metabisulfite, calcium chloride, magnesium chloride, potassium nitrate, and ammonium sulfate.

[0030] The fermented beer-flavored beverage according to this embodiment may be an alcoholic beverage with an alcohol content of 1% v / v or more, or a non-alcoholic beverage with an alcohol content of less than 1% v / v. Unless otherwise specified, "alcohol" in this specification refers to ethanol.

[0031] The alcohol content of the alcoholic beverage is not particularly limited and may be, for example, 1 v / v% or more, 1.5 v / v% or more, 2 v / v% or more, 2.5 v / v% or more, 3 v / v% or more, 3.5 v / v% or more, 4 v / v% or more, 4.5 v / v% or more, or 5 v / v% or more. The alcohol content of the alcoholic beverage may be, for example, 20 v / v% or less, 15 v / v% or less, 10 v / v% or less, 9 v / v% or less, 8 v / v% or less, 7 v / v% or less, 6 v / v% or less, 5 v / v% or less, 4 v / v% or less, 3.5 v / v% or less, 3 v / v% or less, 2.5 v / v% or less, 2 v / v% or less, or 1.5 v / v% or less.

[0032] The alcohol content of a non-alcoholic beverage may be less than 1 v / v%, for example, 0.95 v / v% or less, 0.9 v / v% or less, 0.85 v / v% or less, 0.8 v / v% or less, 0.75 v / v% or less, 0.7 v / v% or less, 0.5 v / v% or less, 0.3 v / v% or less, 0.1 v / v% or less, or less than 0.005 v / v% (0.00 v / v%). The alcohol content of the non-alcoholic beverage may be, for example, 0.005 v / v% or more, 0.01 v / v% or more, 0.05 v / v% or more, 0.1 v / v% or more, 0.2 v / v% or more, 0.3 v / v% or more, 0.4 v / v% or more, 0.45 v / v% or more, 0.5 v / v% or more, 0.55 v / v% or more, 0.6 v / v% or more, 0.65 v / v% or more, or 0.7 v / v% or more.

[0033] The alcohol content of fermented beer-taste beverages can be measured, for example, by the method described in "8.3.6 Beer, Alcohol (Alcolyzer Method)" or "8.3.7 Headspace GC-FID Method" of the Revised BCOJ Beer Analysis Methods (published by the Brewery Association of Japan, edited by the International Technical Committee of Brewers Association (Analysis Committee), expanded and revised in 2013), or by the method described in "3-4 Alcohol Content" of the National Tax Agency's Prescribed Analysis Methods. The alcohol content of fermented beer-taste beverages can be adjusted, for example, by adding alcohol (e.g., raw material alcohol, distilled alcohol such as spirits and vodka, or fermented liquid obtained by brewing), adjusting fermentation conditions, etc.

[0034] The fermented beer-taste beverage according to this embodiment may be either non-sparkling or sparkling. Here, non-sparkling refers to a beverage that has a gas pressure of 0.049 MPa (0.5 kgf / cm) at 20°C. 2 ) and foaming refers to a gas pressure at 20°C of less than 0.049 MPa (0.5 kgf / cm 2 ) or more. In the case of foaming, the upper limit of gas pressure is 0.294 MPa (3.0 kgf / cm 2 ) may be sufficient.

[0035] The fermented beer-flavored beverage according to this embodiment can be served in a container. Any container that can be sealed can be used, and metal (such as aluminum or steel) cans or barrels can be used. Glass containers, PET bottles, paper containers, pouches, and other containers can also be used. There are no particular limitations on the container capacity, and any currently available containers can be used. Metal containers are preferred because they completely block gases, moisture, and light, and can maintain stable quality at room temperature for long periods of time.

[0036] The fermented beer-flavored beverage of this embodiment can be produced according to conventional methods, except for adjusting the apparent fermentation degree to 50% or less and suppressing the production of precursor compounds that are converted into substances that cause aged odor.

[0037] A production method according to one embodiment comprises at least a brewing step and a fermentation step. The production method may further comprise a post-fermentation step. The brewing step is a step of obtaining a pre-fermentation liquid. The fermentation step is a step of fermenting the raw material liquid with yeast. The post-fermentation step is a step of carrying out various post-fermentation treatments (e.g., filtration, sterilization, addition of various additives (e.g., colorants, sweeteners, high-intensity sweeteners, antioxidants, acidulants, flavorings, salts) and the like).

[0038] In the mashing process, a pre-fermentation liquid is obtained using raw materials and mashing water (water used in the mashing process). In other words, the mashing process is a process for preparing a pre-fermentation liquid to be used for fermentation. The mashing process may include, in this order, a saccharification process for producing mash from the raw materials and mashing water, a filtration process for filtering the mash to obtain a sugar-containing liquid, a boiling process for boiling the sugar-containing liquid, a removal process for removing solids from the raw material liquid, and a cooling process for cooling the raw material liquid.

[0039] The saccharification process includes a step of adding raw materials and brewing water, adjusting the temperature to 50 to 76°C, and maintaining that temperature. In this step, the temperature is maintained at 50 to 76°C for, for example, 1 to 200 minutes. This allows, for example, saccharification of the raw materials to proceed and soluble components to elute, resulting in mash containing components necessary for yeast metabolism. The mash obtained in the saccharification process is filtered in a filtration process to produce a sugar-containing liquid.

[0040] In the boiling process, the sugar-containing liquid is boiled to obtain a boiled liquid (a sugar-containing liquid after boiling). A sugar-containing liquid is one that contains components that can be fermented into alcohol by yeast. Examples of sugar-containing liquids include wort and syrup. Wort is an unfermented liquid obtained through the saccharification of the above-mentioned barley raw material. Wort can be obtained, for example, through a process of mixing a raw material such as the above-mentioned barley raw material with water, a process of saccharifying the liquid containing the raw material and water by a conventional method to obtain a saccharified liquid, and a process of filtering the saccharified liquid.

[0041] In the removal step, solids in the post-boiling liquid are removed to obtain a purified liquid. The removal step can be carried out, for example, by precipitating insoluble solids contained in the post-boiling liquid. Examples of solids include thermal coagulation produced in the boiling step. The removal step may be carried out in a whirlpool. In the cooling step, the purified liquid is cooled to a temperature at which fermentation by yeast is possible to obtain a pre-fermentation liquid.

[0042] The fermentation process is a process in which the pre-fermentation liquid is fermented with yeast. The fermentation process results in a post-fermentation liquid obtained by fermenting the pre-fermentation liquid with yeast. In the fermentation process, alcoholic fermentation is carried out with yeast. More specifically, yeast is inoculated into the pre-fermentation liquid and fermented to obtain a post-fermentation liquid containing alcohol produced by the yeast.

[0043] Hops may be added in the method for producing a fermented beer-taste beverage according to this embodiment. Hops may be added during the brewing process, the fermentation process, or the post-fermentation process following the fermentation process, or may be added multiple times. Methods for adding hops include, but are not limited to, kettle hopping, late hopping, and dry hopping. Kettle hopping refers to adding hops while the pre-fermentation liquid is being heated or at the beginning of boiling, while late hopping refers to adding hops just before the end of boiling. Dry hopping refers to adding hops after the start of the fermentation process.

[0044] The production method according to this embodiment may include, as post-fermentation steps, steps of maturing and cooling the post-fermentation liquid, and steps of filtering the post-fermentation liquid. By carrying out the filtration step, insoluble solids, yeast, etc. can be removed from the post-fermentation liquid.

[0045] In the production method according to this embodiment, as another post-fermentation step, the post-fermentation liquid (or the post-fermentation liquid after the filtration step) may be heated (sterilized) or the like.

[0046] The apparent degree of fermentation can be adjusted by adjusting the type and amount of raw materials used, adjusting mashing conditions (e.g., saccharification temperature, saccharification time, use of enzyme preparations), adjusting fermentation conditions (e.g., fermentation temperature, fermentation time, composition of fermentation raw materials), selecting a yeast species with limited sugar assimilation ability, etc. Examples of yeast species with limited sugar assimilation ability include yeast species that do not assimilate maltose and / or maltotriose. Examples of such yeast species include Saccharomycodes ludwigii, Pichia kluyveri, and Torulaspora Delbrueckii. These yeasts may be commercially available and used.

[0047] One way to suppress the production of precursor compounds that are converted into substances that cause aged odor is to inhibit the activity of LOX-1 in the raw material. Methods for inhibiting LOX-1 activity include, for example, inhibiting or blocking LOX-1 activity in the raw material, using a raw material that does not have LOX-1 activity, and combinations thereof.

[0048] Methods for inhibiting or preventing LOX-1 activity in raw materials include, for example, contacting malt with high-temperature water to inactivate LOX-1 in the malt and then dissolving it in the high-temperature water, adjusting the temperature and pH conditions of the saccharification process to reduce LOX-1 activity, and adding a substance that inhibits LOX-1 activity (LOX-1 inhibitor) to the raw materials. An example of a LOX-1 inhibitor is eicosatetraenoic acid.

[0049] An example of a method using a raw material lacking LOX-1 activity is a method using a LOX-1-deficient barley raw material as the barley raw material. As used herein, "LOX-1-deficient barley raw material" refers to a barley raw material lacking LOX-1. As used herein, "LOX-1-deficient" refers to a barley raw material that is substantially free of LOX-1 protein, or that contains LOX-1 protein but is substantially free of LOX-1 activity. LOX-1-deficient barley raw material can be selected, for example, by measuring and confirming the expression level of LOX-1 protein or LOX-1 activity. The LOX-1-deficient barley raw material may be a barley raw material that is itself LOX-1-deficient when used in the production of a beer-flavored beverage. LOX-1-deficient barley raw material can be obtained, for example, by using barley lacking LOX-1 (hereinafter also referred to as "LOX-1-deficient barley") as the barley in a known method for producing a barley raw material. In this case, it is easy to obtain a LOX-1-deficient barley raw material simply by replacing barley with LOX-1-deficient barley.

[0050] LOX-1-deficient barley can be selected, for example, by measuring the expression level of LOX-1 protein or LOX-1 activity. Alternatively, barley having a barley LOX-1 mutant gene, as described below, may be used as the LOX-1-deficient barley.

[0051] The LOX-1 activity of barley can be confirmed, for example, by measuring the linoleic acid oxidation activity of a crude enzyme solution extracted from barley seeds or malt. Specifically, it can be measured, for example, by the following method. One fully ripe barley seed was crushed and extracted with 500 μL of extraction buffer (0.1 M sodium acetate buffer (pH 5.5)) by shaking at 4°C for 30 minutes. The resulting extract was centrifuged at 15,000 rpm for 10 minutes, and the supernatant was collected as a crude enzyme solution. Next, 5 μL of substrate solution (40 mM linoleic acid, 1.0% (W / V) Tween 20 aqueous solution) and 85 μL of extraction buffer were added to 10 μL of the crude enzyme solution, mixed, and then reacted at 24°C for 5 minutes. The reaction was stopped by adding 100 μL of reaction stop solution (80 mM 2,6-di-t-butyl-p-cresol methanol solution) and mixing. The reaction solution was left to stand at -20°C for 30 minutes, then centrifuged at 3,000 rpm for 20 minutes to collect the supernatant. To 20 μL of the resulting supernatant, 200 μL of color development solution (4 mM 2,6-di-t-butyl-p-cresol, 25 mM sulfuric acid, 0.25 mM ammonium iron(II) sulfate hexahydrate, 100 mM xylenol orange, 90% aqueous methanol) was added, and after 30 minutes of incubation, the absorbance at 550 nm was measured. As a negative control, the crude enzyme solution was heat-treated at 100°C for 5 minutes to inactivate LOX-1, and then reacted in the same manner. As a positive control, a crude enzyme solution from seeds of the barley variety Kendall was used.

[0052] The expression level of barley LOX-1 protein can be examined by, for example, Western blotting using an anti-LOX-1 antibody, specifically, by the following method. Three micrograms of total soluble protein extracted from barley using 0.1 M sodium acetate buffer (pH 5.5) was fractionated by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and then blotted onto a PVDF membrane (Millipore). The membrane was washed with TTBS (20 mM Tris-HCl (pH 7.5), 0.15 M sodium chloride, 0.05% (w / v) Tween 20, 0.05% (w / v) sodium azide) and then incubated with LOX-1 antibody solution (1:1000 dilution in TTBS) for 30 minutes. The membrane was washed three times for 5 minutes with TTBS and then incubated with alkaline phosphatase-conjugated goat anti-rabbit IgG antibody solution (Santa Cruz, 1:1000 dilution in TTBS) for 30 minutes. The membrane is washed twice for 5 minutes with TTBS, and once for 5 minutes with AP9.5 (10 mM Tris-HCl (pH 9.5), 0.1 M sodium chloride, 5 mM magnesium chloride), and then reacted with alkaline phosphatase substrate solution (1 mg / ml nitroblue tetrazolium, 0.5 mg / ml BCIP, AP9.5 solution) to develop color. This allows comparison of expression levels based on the intensity of the bands relative to the control.

[0053] LOX-1-deficient barley is, for example, barley having a mutant barley LOX-1 gene in which guanine in the splicing donor site (5'-GT-3') of the fifth intron of the barley LOX-1 gene has been mutated to adenine. This mutant gene is characterized by a substitution of base G at position 60 in the fifth intron of the LOX-1 gene with A, as compared with known LOX-1 genes. Because positions 60-61 of the fifth intron of the LOX-1 gene are the splicing donor site (5'-GT-3'), this substitution of bases causes abnormal splicing of LOX-1, preventing the expression of active LOX-1.

[0054] As described above, LOX-1-deficient barley is characterized by not expressing LOX-1 protein or having substantially no LOX-1 activity, and preferably also has the characteristics of a normal barley variety suitable for producing beer-taste beverages, the characteristics of a barley variety suitable for breeding, etc. LOX-1-deficient barley having the desired characteristics can be obtained, for example, by crossing LOX-1-deficient barley carrying a mutant LOX-1 gene with a variety having those characteristics.

[0055] LOX-1-deficient barley raw material can also be obtained by malting normal barley, i.e., barley that is not LOX-1-deficient, using a known method for reducing LOX-1. [Example]

[0056] The present invention will be described in more detail below with reference to examples, although the present invention is not limited to the following examples.

[0057] Test Example 1: Production of fermented beer-flavored beverage Malt was obtained by malting LOX-1-deficient barley or conventional barley (barley with LOX-1 activity). LOX-1-deficient barley has a mutant LOX-1 gene in which the guanine in the splicing donor site (5'-GT-3') of the fifth intron of the barley LOX-1 gene is mutated to adenine.

[0058] The yeast used in this study was either bottom-fermenting yeast (Saccharomyces pastorianus) or brewer's yeast (Saccharomycodes ludwigii), which has low alcohol production. S. ludwigii utilizes glucose, sucrose, and fructose, but does not utilize maltose or maltotriose, resulting in little alcohol production.

[0059] Using brewing water, malt, and hops (malt usage ratio 100% by mass), saccharification, boiling, solids removal, and cooling were carried out according to standard methods to produce chilled wort (brewing step). The resulting chilled wort was then inoculated with yeast, and alcoholic fermentation was carried out according to standard methods (fermentation step). The resulting post-fermentation liquid was then filtered, and the gas pressure was adjusted to obtain the fermented beer-flavored beverages of Test Examples 3 and 4.

[0060] After carrying out the mashing and fermentation steps in the same manner as in Test Examples 3 and 4, an acidulant was added to the post-fermentation liquid. The resulting post-fermentation liquid was then filtered, and the gas pressure was adjusted to obtain the fermented beer-taste beverages of Test Examples 1 and 2.

[0061] Except for using brewing water, malt, cornstarch, and hops (malt usage ratio: 51% by mass), the brewing and fermentation processes were carried out in the same manner as in Test Examples 3 and 4, and then an acidulant was added to the post-fermentation liquid. The resulting post-fermentation liquid was then filtered, and the gas pressure was adjusted to obtain the fermented beer-taste beverages of Test Examples 5 and 6.

[0062] The types of malt and yeast used in the fermented beer-taste beverages of Test Examples 1 to 6 are as shown in Table 1.

[0063] The extract of the cold wort (original extract), and the apparent extract, apparent fermentation degree, true extract, alcohol content, and pH of the fermented beer-flavored beverage were measured or calculated according to the method described in the Revised BCOJ Beer Analysis Methods (published by the Brewery Society of Japan, edited by the International Technical Committee of the Brewers Association of Japan (Analysis Committee), revised and expanded in 2013). The results are also shown in Table 1.

[0064] [Table 1]

[0065] Test Example 2: Evaluation of antioxidant activity of fermented beer-flavored beverages The antioxidant activity of the resulting fermented beer-flavored beverages of Test Examples 1 to 4 was evaluated by the ORAC (Oxygen Radical Absorbance Capacity Assay) method. Specifically, according to the method described in a non-patent document (J. Agric. Food Chem., 2004, Vol. 52, No. 12, pp. 4026-4037), a radical donor (2,2'-azobis(2-amidinopropane)dihydroxychloride), a fluorescent substance (fluorescein), and a sample or standard (Trolox) were mixed, and the fluorescence intensity was measured over time. The area enclosed by the curve of fluorescence intensity versus reaction time when the sample or standard was added and the curve of fluorescence intensity versus reaction time when the sample or standard was not added was calculated as the antioxidant activity. The antioxidant activity of each sample was expressed as a relative value per unit weight, with the antioxidant activity exhibited by 1 μmol of standard (Trolox) defined as 1 μmol TE. The results are shown in Table 2.

[0066] [Table 2]

[0067] The fermented beer-flavored beverages of Test Examples 1 and 2, obtained by fermentation with S. ludwigii, had antioxidant activity below the detection limit (5 μmol TE / g). This indicates that the fermented beer-flavored beverages with a low degree of fermentation obtained by fermentation with S. ludwigii have significantly weaker antioxidant power.

[0068] Test Example 3: Evaluation of sulfite concentration in fermented beer-flavored beverages The sulfite concentrations in the fermented beer-flavored beverages obtained in Test Examples 1 to 6 were measured using the method (titration) described in the Revised BCOJ Beer Analysis Methods (published by the Brewery Society of Japan, edited by the International Technical Committee of the Brewers Association of Japan (Analysis Committee), revised and expanded in 2013). The results are shown in Table 3.

[0069] [Table 3]

[0070] The fermented beer-flavored beverages of Test Examples 1-2 and 5-6 obtained by fermentation with S. ludwigi had concentrations of sulfite, an antioxidant, below the detection limit (0.3 mg / L).

[0071] Test Example 4: Evaluation of flavor durability of fermented beer-flavored beverage (1) The resulting fermented beer-taste beverages of Test Examples 1 to 6 were stored for one month at 5° C., 20° C., or 30° C. Thereafter, the degree of staling of the fermented beer-taste beverages of Test Examples 1 to 6 was evaluated by sensory evaluation.

[0072] The sensory evaluation was carried out by seven selected panelists with distinguishing abilities. The degree of aging, which combined the aging odor and aging taste, was evaluated on a nine-point scale from 0 to 4 (in increments of 0.5 points), and the average score was used as the evaluation score. A higher score indicates a higher degree of aging. The results are shown in Table 4.

[0073] [Table 4]

[0074] The fermented beer-taste beverages of Test Examples 3 and 4, which have a high degree of attenuation, had a low degree of staling after one month of storage, regardless of the type of malt, due to their high antioxidant capacity, as confirmed in Test Examples 2 and 3. The fermented beer-taste beverages of Test Examples 1-2 and 5-6, which have a low degree of attenuation, had a low antioxidant capacity and therefore staling progressed quickly, as confirmed in Test Examples 2 and 3, but the degree of staling was significantly improved by using malt derived from LOX-1-deficient barley. Furthermore, even among the fermented beer-taste beverages with a low degree of attenuation, the fermented beer-taste beverages of Test Examples 5 and 6, which use a lower proportion of malt, were found to have a lower degree of staling and higher flavor durability.

[0075] Test Example 5: Evaluation of flavor durability of fermented beer-flavored beverages (2) The resulting fermented beer-taste beverages of Test Examples 1 to 6 were stored for three months at 5° C., 20° C., or 30° C. The trans-2-nonenal concentrations in the fermented beer-taste beverages of Test Examples 1 to 6 were measured immediately after production and one month, two months, and three months after the start of storage.

[0076] The trans-2-nonenal concentration in the fermented beer-flavored beverage was measured by solid-phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS) using the standard addition method. The results are shown in Tables 5 to 7.

[0077] [Table 5]

[0078] [Table 6]

[0079] [Table 7]

[0080] As shown in Tables 5 to 7, the use of malt derived from LOX-1-deficient barley reduced the trans-2-nonenal concentration (comparison between Test Example 1 and Test Example 2, comparison between Test Example 3 and Test Example 4, and comparison between Test Example 5 and Test Example 6). In the fermented beer-taste beverages of Test Examples 3 and 4, which had a high degree of attenuation, the trans-2-nonenal concentration was low even when malt derived from normal barley was used, and the improvement effect of using malt derived from LOX-1-deficient barley was small. On the other hand, in the fermented beer-taste beverages of Test Examples 1-2 and 5-6, which had a low degree of attenuation, the use of malt derived from LOX-1-deficient barley significantly reduced the trans-2-nonenal concentration, demonstrating a significant improvement effect. Furthermore, even among the fermented beer-taste beverages with a low degree of attenuation, the fermented beer-taste beverages of Test Examples 5 and 6, which used a lower proportion of malt, showed a lower trans-2-nonenal concentration and higher flavor durability.

[0081] LOX-1, an enzyme contained in barley, is known to oxidize linoleic acid to produce 9-hydroperoxyoctadecadienoic acid. 9-Hydroperoxyoctadecadienoic acid is also known to be converted to trans-2-nonenal, one of the substances that causes aged odor. The results of Test Examples 4 and 5 demonstrate that the flavor durability of fermented beer-flavored beverages with a low degree of attenuation can be improved by inhibiting or blocking the activity of LOX-1.

Claims

1. The apparent degree of fermentation is 50% or less, A fermented beer-flavored beverage in which the increase in trans-2-nonenal concentration when stored at 20°C for one month is 0.1 μg / L or less.

2. 2. The fermented beer-taste beverage according to claim 1, wherein the increase is 0.05 μg / L or less.

3. The apparent degree of fermentation is 50% or less, A fermented beer-flavored beverage having a trans-2-nonenal concentration of 0.2 μg / L or less when stored at 20°C for one month or more.

4. 4. The fermented beer-taste beverage according to claim 1 or 3, having an ORAC of 5 μmol TE / g or less.

5. 4. The fermented beer-taste beverage according to claim 1 or 3, wherein the sulfite concentration is 5 mg / L or less.

6. 4. The fermented beer-taste beverage according to claim 1 or 3, having an alcohol content of less than 1 v / v %.

7. 4. The fermented beer-taste beverage according to claim 1 or 3, having an apparent degree of fermentation of 40% or less.

8. 4. The fermented beer-taste beverage according to claim 1 or 3, wherein the malt content is 30% by mass or more and less than 100% by mass.

9. The apparent degree of fermentation is 50% or less, A fermented beer-flavored beverage having an ORAC of 5 μmol TE / g or less.

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