JP7136852B2 - Sugar-free beer-taste fermented alcoholic beverage - Google Patents

Description

The present invention relates to a sugar-free, beer-taste fermented alcoholic beverage, more specifically, a sugar obtained by adjusting the total concentration of peptides having a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis) and isomaltose and panose to a predetermined concentration. It relates to a quality zero beer-taste fermented alcoholic beverage.

Due to the recent increase in health consciousness, there is a demand for beer-taste beverages with reduced sugar content. Various techniques have been developed so far with the aim of producing beer-taste beverages with reduced sugar content.

For example, as a technique for stably producing a beer-taste fermented alcoholic beverage containing no carbohydrates, a technique using assimilable monosaccharides and assimilable disaccharides as sugar sources (Patent Document 1) is known. In addition, as a technique for producing sugar-free beer-taste beverages by a method with extremely low risk of microbial contamination, a technique is known in which the content of oligosaccharides in carbohydrate raw materials is used within a specific range (Patent Document 2). . In addition, there is known a technique (Patent Document 3) of realizing a rich taste in beer-taste fermented alcoholic beverages by adjusting the ratio of peptides with a molecular weight of 800 to 1500 Da in the beverage within a specific range.

Japanese Patent Application Laid-Open No. 2009-131202 JP 2012-157323 A JP 2019-154438 A

Among beer-taste fermented alcoholic beverages with reduced sugar content, so-called “zero sugar” beer-taste fermented alcoholic beverages with a sugar concentration of less than 0.5 g/100 mL have a considerable amount of sugar reduced. In particular, it gives a watery and thin flavor impression. In Patent Document 3, the depth of taste can be imparted by adjusting the ratio of peptides with specific molecular weights, but on the other hand, the present inventors have found that as the peptide concentration increases, the off-taste also tends to increase. I came to recognize the problem.

That is, an object of the present invention is to provide a sugar-free beer-taste fermented alcoholic beverage that has a rich flavor and reduced unpleasant taste.

The present inventors have found that in a beer-taste fermented alcoholic beverage in which the sugar concentration is reduced to less than 0.5 g/100 mL, the concentration of peptides with a molecular weight of 800 to 1500 Da and the concentration of non-utilizable sugars are adjusted within a specific range. , found that the beer-taste fermented alcoholic beverage can achieve a rich taste while reducing the unpleasant taste caused by the peptide. The present invention is based on this finding.

According to the present invention, the following inventions are provided.
[1] A beer-taste fermented alcoholic beverage containing at least a portion of malt and/or ungerminated barley and having a carbohydrate concentration of less than 0.5 g/100 mL, wherein the beverage has a molecular weight of 800 to 1500 Da ( A beer-taste fermented alcoholic beverage having a peptide concentration of 0.136 mg/mL or more (by gel filtration method for HPLC analysis) and a total concentration of isomaltose and panose of 0.01 to 0.15 g/100 mL.
[2] The beer-taste fermented alcoholic beverage according to [1] above, wherein the malt content is 50% or more.
[3] A method for producing a beer-taste fermented alcoholic beverage having a sugar concentration of less than 0.5 g/100 mL, which uses malt and/or ungerminated barley as at least a part of raw materials and has improved flavor, Adjust the concentration of peptides with a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis) in the beverage to 0.136 mg / mL or more, and the total concentration of isomaltose and panose to 0.01 to 0.15 g / 100 mL A manufacturing method comprising conditioning.
[4] In the production process of beer-taste fermented alcoholic beverages, the molecular weight of 800 to 1500 Da contained in beer-taste fermented alcoholic beverages containing at least part of malt and/or ungerminated barley as raw materials (gel filtration method for HPLC analysis) and/or adding isomaltose and/or panose, the production method according to [3] above.
[5] A method for improving the flavor of a beer-taste fermented alcoholic beverage having a sugar concentration of less than 0.5 g/100 mL, wherein at least part of the raw material is malt and/or ungerminated barley, wherein the molecular weight in the beverage is Adjusting the peptide concentration of 800 to 1500 Da (gel filtration method for HPLC analysis) to 0.136 mg / mL or more, and adjusting the total concentration of isomaltose and panose to 0.01 to 0.15 g / 100 mL How to be.

According to the present invention, in a beer-taste fermented alcoholic beverage having a sugar concentration of less than 0.5 g/100 mL, the concentration of peptides having a molecular weight of 800 to 1500 Da and the concentration of non-utilizable sugars are adjusted within specific ranges, It is possible to provide a sugar-free, beer-taste fermented alcoholic beverage that achieves a rich taste while reducing the unpleasant taste derived from.

FIG. 2 is a diagram showing a calibration curve created from the retention time of the gel filtration method for HPLC analysis and the molecular weight of known substances performed in Reference Example 2. FIG. 2 is a diagram showing sensory evaluation results of Example 1. FIG. The effect of the addition of peptide fractions with a molecular weight of 800 to 1500 Da and the addition of non-utilizable sugars on the depth of taste and off-taste was shown by sensory evaluation scores.

Specific description of the invention

In the present invention, the "beer taste" means the taste and aroma peculiar to beer obtained when beer is normally produced, that is, when beer is produced based on fermentation with yeast or the like.

In the present invention, the "beer-taste fermented alcoholic beverage" is a beer-taste fermented alcoholic beverage fermented with yeast using a carbon source, a nitrogen source, water, etc. as raw materials, and uses beer, low-malt beer and malt as raw materials. Beverages obtained by adding alcohol to beer or low-malt beer (for example, liqueur-based new-genre beverages classified as “liqueur (sparkling) (1)” under the Liquor Tax Act).

The beer-taste fermented alcoholic beverage of the present invention uses malt and/or ungerminated barley as part of the raw material, and can take any malt use ratio, but for example, the malt use ratio is 25. %, 25% or more, less than 50%, 50% or more, 60% or less, 70% or less, 80% or less, or 95% or less, and these upper and lower limits can be arbitrarily combined. . As used herein, the term "proportion of malt used" refers to the ratio of malt mass to the mass of all raw materials excluding brewing water.

The beer-taste fermented alcoholic beverage of the present invention is a sugar-free beverage with a reduced sugar concentration. Specifically, the sugar concentration of the beer-taste fermented alcoholic beverage of the present invention is less than 0.5 g/100 mL. is.

The sugar concentration can be measured by a known method, and a method of calculating the amount of water, protein, lipid, ash and dietary fiber from the mass of the sample (Nutrition Labeling Standards (December 16, 2009) This can be done in accordance with Consumer Affairs Agency Notification No. 9 (partially revised)).

In the beer-taste fermented alcoholic beverage of the present invention, the lower limit (or more or more) of the concentration of peptides with a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis) in the beverage is 0.136 mg/mL, 0.14 mg/mL, 0. .15 mg/mL, 0.20 mg/mL or 0.30 mg/mL, and an upper limit (less than or equal to) of 2.50 mg/mL, 2.00 mg/mL, 1.50 mg/mL, It can be 1.20 mg/mL, 1.00 mg/mL, 0.70 mg/mL, 0.65 mg/mL or 0.60 mg/mL. These lower and upper limits can be combined arbitrarily, and the concentration of peptides with a molecular weight of 800-1500 Da (HPLC analysis gel filtration method) in the beverage is, for example, 0.136-2.50 mg/mL, 0.136-2.50 mg/mL, It can be 20-2.00 mg/mL, 0.20-1.50 mg/mL or 0.30-1.50 mg/mL.

In the beer-taste fermented alcoholic beverage of the present invention, the ratio of the mass of peptides with a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis) in the beverage to the total peptide mass derived from the beverage fractionated by the gel filtration method for HPLC analysis. (percentage) can be within a specific range. The ratio is the total peptide concentration (mg/mL) derived from the beverage fractionated by the HPLC analytical gel filtration method from the peptide concentration (mg/mL) with a molecular weight of 800 to 1500 Da (HPLC analytical gel filtration method) in the beverage. It can be calculated by dividing by In the present invention, the lower limit (or more or more) of the peptide ratio can be 15%, 16% or 17% from the viewpoint of imparting depth of taste, and the upper limit (less than or equal to or less than) is It can be 45%, 35% or 25%. These lower and upper limits can be arbitrarily combined, for example, the peptide ratio range is 15 to 45%, 16 to 45%, 17 to 45%, 17 to 25%, or 17% to less than 25%. can be

In the beer-taste fermented alcoholic beverage of the present invention, the peptide concentration (mg/mL) with a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis) in the beverage relative to the original extract concentration (OE concentration) (°P) in the beverage The lower limit (or more or more) of the ratio of can be set to 0.02 (preferably 0.03), and the upper limit (less than or equal to) is 0.50 (preferably 0.30, 0.30, 0.50). 15, 0.12, 0.10 or 0.08). These lower and upper limits can be combined arbitrarily, for example, peptide concentration (mg / mL ) can be 0.02 to 0.50 (preferably 0.03 to 0.30).

The lower limit (or more or more) of the OE concentration in the beer-taste fermented alcoholic beverage of the present invention can be 2°P (preferably 4°P), and the upper limit (below or less than) is 20°P. (preferably 14°P, more preferably 10°P). These lower and upper limits can be combined arbitrarily. For example, the OE concentration in the beer-taste fermented alcoholic beverage of the present invention can be 2 to 20°P (preferably 4 to 14°P). . In the present invention, the "original extract concentration" can be measured using a commercially available device (for example, alcoholizer (manufactured by Anton Paar)). In the case of beer-taste fermented alcoholic beverages at least partly made of malt and/or ungerminated barley, the “original extract concentration” can be replaced with the “original wort extract concentration”.

The molecular weight of peptides in beverages is measured by a gel filtration method using high-performance liquid chromatography (herein sometimes referred to as "HPLC analysis gel filtration method"). Specifically, the molecular weight can be calculated from the retention time using a gel filtration calibration curve for HPLC analysis (for example, FIG. 1), and samples having different molecular weights depending on the retention time can be fractionated. A specific example of molecular weight measurement by gel filtration for HPLC analysis is as shown in Reference Example 2 below. Also, the peptide can be quantified by the Lowry method.

In the beer-taste fermented alcoholic beverage of the present invention, the lower limit (or more or more) of the total concentration of isomaltose and panose in the beverage is 0.01 g/100 mL, 0.02 g/100 mL, 0.03 g/100 mL, 0.03 g/100 mL, 04 g/100 mL, 0.05 g/100 mL, and the upper limit (or less or less) is 0.15 g/100 mL, 0.14 g/100 mL, 0.13 g/100 mL, 0.12 g/100 mL, 0 .11 g/100 mL, 0.10 g/100 mL. These lower and upper limits can be combined arbitrarily. 0.02-0.15 g/100 mL or 0.02-0.10 g/100 mL. The total concentration of isomaltose and panose can be calculated by adding the concentrations of isomaltose and panose in the beverage.

The concentrations of isomaltose and panose in beverages can be measured by ion chromatography.

The lower limit (or more or more) of the alcohol content in the beer-taste fermented alcoholic beverage of the present invention can be 1 v/v%, 2 v/v% or 3 v/v%, and the upper limit (below or less than ) can be 9 v/v %, 8 v/v % or 7 v/v %. These lower and upper limits can be combined arbitrarily, for example, the alcohol content in the beverage can be 1 to 9 v/v%, 2 to 8 v/v%, and 3 to 7 v/v%. .

Although the beer-taste fermented alcoholic beverage of the present invention has a reduced sugar concentration, it is imparted with a rich taste and has a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis). It is characterized by reduced taste. As used herein, the term "thickness of taste" means a flavor sensation recognized by the breadth of taste, complexity, and imparting a body feeling. "Miscellaneous taste" means astringent taste, harsh taste and astringent taste.

In the beer-taste fermented alcoholic beverage of the present invention, the sugar concentration in the beverage is reduced within a predetermined range, and the peptide concentration of 800 to 1500 Da and the total concentration of isomaltose and panose in the beverage are each within a predetermined range. As long as it is adjusted to , it can be produced according to the normal procedure for producing beer-taste fermented alcoholic beverages.

For example, fermentation is performed by adding fermentation brewer's yeast to wort prepared from brewing raw materials such as malt, hops, auxiliary raw materials, and brewing water, and the resulting fermented liquid is stored at a low temperature and then filtered. A beer-taste fermented alcoholic beverage can be produced by removing the yeast. Of course, among beer-taste fermented alcoholic beverages, all-malt beer can be produced from malt, hops, and water.

In the production procedure described above, wort can be prepared according to a conventional method. For example, preparing wort by saccharifying a mixture of brewing raw materials and brewing water, filtering to obtain wort, adding hops to the wort, boiling, and cooling the boiled wort. can be done. Also, the wort can be prepared by adding a commercially available enzyme preparation during the saccharification process. For example, protease preparations for protein degradation, α-amylase preparations, glucoamylase preparations, pullulanase preparations for carbohydrate degradation, β-glucanase preparations, fibrinolytic enzyme preparations for fibrinolysis, etc. can be used, or a mixed formulation thereof can also be used.

In one aspect of the production of the beer-taste fermented alcoholic beverage of the present invention, malt with high endo-type protease activity and/or enzyme preparations with endo-type protease activity are used for proteolysis in the saccharification step or fermentation step. It is also possible to extend the optimal saccharification step time of endo-type proteases. By carrying out such a step, the concentration of peptides with a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis) contained in the beer-taste fermented alcoholic beverage can be increased and adjusted within a predetermined value range. It is possible to produce a beer-taste fermented alcoholic beverage with a rich taste while containing zero sugar. That is, according to the present invention, there is provided a method for producing a beer-taste beverage comprising using an enzyme preparation containing malt and/or an endoprotease with high endoprotease activity in a saccharification step or a fermentation step. The characteristics of the enzymatic activity of the above malt and enzyme preparation can be specified based on the endo-type protease activity.

In one aspect of the production of the beer-taste fermented alcoholic beverage of the present invention, in the saccharification step, a glucoamylase enzyme preparation or an enzyme preparation having glucoamylase activity can be used to decompose carbohydrates, and the saccharification time is By performing all or part of these, the concentration of isomaltose and panose contained in the beer-taste fermented alcoholic beverage is increased, and the concentration is within a predetermined value range fermented alcoholic beverage with a beer taste, which can be adjusted within the range of 0.5 to 100 nm, further enhancing the depth of taste imparted by a peptide having a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis), and reducing the unfavorable taste caused by the peptide. can be manufactured.

The beer-taste fermented alcoholic beverage of the present invention uses at least one or both of malt and ungerminated barley as a raw material, preferably malt or malt and ungerminated barley as a raw material. It is intended to be a part. In the beer-taste fermented alcoholic beverage of the present invention, barley malt can be used as part of the malt. In the beer-taste fermented alcoholic beverage of the present invention, ungerminated barley (including extract) and ungerminated wheat (including extract) are used as raw materials. can be done.

In the production of the beer-taste fermented alcoholic beverage of the present invention, in addition to malt and ungerminated barley, rice, corn, soybeans, koryan, potatoes, starch, sugars (e.g. liquid sugar), fruits (e.g. fruit juice, concentrated fruit juice), coriander or its seeds, spices or their raw materials (e.g. pepper, cinnamon, sansho), herbs (e.g. chamomile, sage, basil), vegetables (e.g. sweet potato, pumpkin), buckwheat or sesame, carbohydrates (e.g., honey, brown sugar), salt, miso, flowers, tea, coffee, cocoa (tea, coffee and cocoa include preparations thereof), marine products (e.g., oysters, kelp, wakame seaweed, dried bonito) and other auxiliary ingredients; Nitrogen sources such as protein hydrolyzate and yeast extract; and other additives such as fragrances, pigments, foaming/foam retention improvers, water conditioners, and fermentation aids can be used as brewing raw materials. In the beer-taste fermented alcoholic beverage of the present invention, at least malt and hops can be used as raw materials other than brewing water, and in some cases, sugars, rice, corn, starch, etc. can also be used as raw materials.

In the present invention, a fraction containing peptides with a molecular weight of 800 to 1500 Da is prepared from cereals (e.g., malt and/or ungerminated barley and soybeans), and the fraction is treated as sugar-free beer-taste fermented alcohol. By adding it to the beverage, the concentration of the peptide having a molecular weight of 800 to 1500 Da is adjusted within a predetermined value range, and the non-utilizable sugar (isomaltose and/or panose) is added to the beer-taste beverage. A beer-taste fermented alcohol capable of adjusting the concentration of assimilable sugars within a predetermined value range and having a rich taste while being zero sugars and having a reduced unpleasant taste caused by the peptide. Beverages can be produced. That is, according to the present invention, a fraction containing peptides with a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis) is prepared from cereals, and the fraction is treated with beer having a sugar concentration of less than 0.5 g/100 mL. A method for producing a beer-taste fermented alcoholic beverage is provided, comprising adding to a taste-fermented alcoholic beverage, and adding a non-utilizable sugar to the beer-taste fermented alcoholic beverage. The addition of the peptide fraction and the non-utilizable sugar to the beer-taste fermented alcoholic beverage can be carried out during the production process of the beverage (for example, the process of adding the fermented liquid). Peptides with a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis) derived from cereals can be prepared, for example, by hydrolyzing cereals with a proteolytic enzyme such as peptidase. It may also be obtained from protein hydrolysates of cereals such as cereals. As the non-utilizable sugars, isomaltose and panose, those obtained from food materials or commercially available products can be used.

In the present invention, a beer-taste fermented alcoholic beverage is produced using at least a portion of malt and/or ungerminated barley as a raw material, and a fraction containing peptides with a molecular weight of 800 to 1500 Da is prepared from the beverage, By adding this fraction to a sugar-free beer-taste fermented alcoholic beverage, the concentration of peptides with a molecular weight of 800 to 1500 Da is adjusted within a predetermined range, and non-utilizable sugars (isomaltose and/or panose ) to the beer-taste beverage, the concentration of the non-utilizable sugars can be adjusted within a predetermined value range, and thus the depth of taste is imparted while the sugar is zero, and the peptide It is possible to produce a beer-taste fermented alcoholic beverage with reduced unpleasant taste. That is, according to the present invention, a beer-taste fermented alcoholic beverage containing at least a portion of malt and/or ungerminated barley as a raw material is produced, and then the beverage has a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis). ), adding the fraction to a beer-taste fermented alcoholic beverage with a sugar concentration of less than 0.5 g / 100 mL, and non-utilizable sugars (isomaltose and / or Panose) to the beer-taste fermented alcoholic beverage is provided. The addition of the peptide fraction and the non-utilizable sugar to the beer-taste fermented alcoholic beverage can be carried out during the production process of the beverage (for example, the process of adding the fermented liquid).

In addition, according to the present invention, one or more peptides having a molecular weight of 800 to 1500 Da derived from beer-taste fermented alcoholic beverages at least partly made of cereals or malt and/or ungerminated barley, and A carbohydrate-free beer-taste fermented alcoholic beverage containing non-utilizable sugars (isomaltose and/or panose) is provided, and the beverage is a part of the beer-taste fermented alcoholic beverage of the present invention. The beverage containing the peptide and the non-utilizable sugar contains no sugar, but has an improved or improved flavor as a beer-taste fermented alcoholic beverage. In addition, the beverage is reduced in off-taste caused by the peptide.

Sugars in beer-taste fermented alcoholic beverages can be reduced according to known methods, for example, a method of adding glucoamylase during the saccharification process, a method of adding glucoamylase during the fermentation process・ From analysis to modification, advanced functionality, and industrial use (see Makoto Komiyama, NTS Co., Ltd., 2010), yeast using liquid sugar containing a lot of sugar that is assimilated by yeast A method of reducing sugar by increasing the assimilation by using (see JP-A-2009-131202), a method of wort filtration during the saccharification process (see JP-A-2012-147780) Sugar in beverages The quality concentration can be set to a predetermined value.

The beverage provided by the present invention may optionally be subjected to a step of adding carbon dioxide gas, and may be provided as a container-packaged beverage through steps such as a filling step and a sterilization step. Sterilization may be before or after filling the container. Further, when the pH of the beverage is adjusted to less than 4, the filling step can be performed as it is without the sterilization step to obtain a packaged beverage.

The container used for the beverage according to the invention may be any container normally used for filling beverages, such as metal cans, barrels, plastic bottles (e.g. PET bottles, cups), paper containers, bottles, Pouch containers and the like can be mentioned, but metal can/barrel containers, plastic bottles (for example, PET bottles), and bottles are preferred.

According to another aspect of the present invention, there is provided a method for producing a sugar-free, beer-taste fermented alcoholic beverage containing at least a portion of malt and/or ungerminated barley with improved flavor, comprising: Adjust the concentration of peptides with a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis) in the beverage to 0.136 mg / mL or more, and the total concentration of isomaltose and panose to 0.01 to 0.15 g / 100 mL A method is provided comprising adjusting. In the present invention, the “improved flavor” or “improved flavor” of a beer-taste fermented alcoholic beverage means that a beer-taste fermented alcoholic beverage with zero carbohydrates has a rich taste and an unfavorable taste caused by the peptide. is reduced. The production method of the present invention described above is a method for producing a sugar-free, beer-taste fermented alcoholic beverage that is preferably imparted with a rich taste and that has a reduced unpleasant taste caused by the peptide. The above production method of the present invention can be carried out according to the description of the beer-taste fermented alcoholic beverage of the present invention and its production method.

According to another aspect of the present invention, there is also provided a method for improving the flavor of a sugar-free, beer-taste fermented alcoholic beverage using malt and/or ungerminated barley as part of the raw material. The method for improving the flavor of the present invention comprises adjusting the concentration of peptides having a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis) in the beverage to 0.136 mg/mL or more, and reducing the total concentration of isomaltose and panose to 0.136 mg/mL. 01-0.15 g/100 mL. The flavor-improving method of the present invention is preferably a method of imparting a depth of taste to a sugar-free, beer-taste fermented alcoholic beverage, and reducing the off-taste caused by the peptide. The method for improving the flavor of the present invention can be carried out according to the description regarding the beer-taste fermented alcoholic beverage of the present invention and the method for producing the same.

The present invention will be described more specifically based on the following examples, but the present invention is not limited to these examples.

Analysis of carbohydrate concentration Measurement of carbohydrate concentration based on the Nutrition Labeling Standards was carried out according to the known method described in the 5th Revised Japanese Food Standard Composition Table Analysis Manual. That is, from the mass of the sample beverage, moisture (heat drying method under reduced pressure), protein (method using an automatic analyzer), lipid (Soxhlet extraction method (3)), ash (combustion method using a muffle furnace), and dietary fiber content (Prosky It was measured by a calculation method excluding the enzyme weight method).

Reference Example 1: Identification of a peptide fraction that imparts a preferable flavor to a beer-taste beverage (1) Production of a beer-taste fermented alcoholic beverage Barley malt, hops, and an enzyme preparation are used to produce a beer-taste fermented alcoholic beverage by an infusion method. manufactured. Specifically, 100 g of barley malt was added to 300 mL of hot water at 65° C., held for 60 minutes, further heated to 78° C., held for 5 minutes, filtered, and wort was obtained. Subsequently, 0.8 g/L of hops were added, and the mixture was boiled at 100° C. for 90 minutes and then filtered to obtain a pre-fermentation liquid.

After that, fermentation was performed with beer yeast according to a conventional method, and the flavor of the fermented liquid was confirmed. As a result, it was confirmed that the obtained fermented liquid had suppressed unpleasant taste, had a beer-like soft and smooth texture, and had a well-balanced taste.

(2) Gel Filtration Fractionation The fermented liquid obtained in (1) above was filtered through a 0.45 μm filter, and the filtered fermented liquid was weighed and freeze-dried. The dried product was dissolved in a 100 mM NaCl solution to prepare a 5-fold concentrated solution, which was used as a sample for fractionation. The sample was subjected to gel filtration fractionation under the following conditions.

<Gel filtration fractionation conditions>
Column: Hiload Superdex 30pg 26/600 (manufactured by GE Healthcare)
Sample injection volume: 5mL
Eluent composition: 100 mM NaCl
Flow rate: 2.5 mL/min (constant flow rate)
Detection wavelength: 215 nm
Prep: 0.29 cv (column volume) to 19.1 mL (fraction 0), then 5 mL fractions from 0.35 cv (fractions 1-51)

Based on the sensory evaluation of the fractions, the fractions were divided into 9 groups of pre-fractions, A1, A2, B, C, D, E, F, and G, as shown in Table 1, according to differences in flavor characteristics. Also, the flavor characteristics of each fraction were determined by discussion of 5 trained panelists.

(3) Purification of Peptide Fraction Fractions C and D obtained in (2) above were subjected to adsorption treatment using a solid-phase extraction column (Bond Elute C18, manufactured by Agilent Technologies) and washed with demineralized water. , eluted with 50% ethanol aqueous solution, concentrated to dryness, and recondensed with desalted water to obtain a concentrated solution. This was used as a peptide fraction.

(4) Protein quantification by the Lowry method The protein quantification of the peptide fraction obtained by purifying the gel filtration fraction in (2) and the peptide fraction in (3) was performed using a commercially available kit (DC protein assay, Bio- Rad Co.) was performed by the Lowry method. First, the concentration of the above-mentioned fractionated liquid was adjusted to an appropriate range. To 5 μL of the concentration-adjusted sample, 50 μL of solution A was added and stirred, and then 400 μL of solution B was added and stirred. After color development reaction was performed at room temperature for 15 minutes, 350 μL of the solution was transferred to a 96-well plate and absorbance at 750 nm was measured. Peptide concentration (mg/mL) was calculated based on the obtained absorbance and a previously prepared calibration curve. A calibration curve was prepared using BSA (bovine serum albumin).

(5) Results Peptide fractions were analyzed with a Superdex 75 10/300 column to estimate the molecular weight, and the peptides of fractions C and D were distributed over a molecular weight range of approximately 800-1500 Da.

From the above results, it was suggested that peptide fractions C and D with a molecular weight of about 800 to 1500 Da fractionated and purified from beer-taste fermented alcoholic beverages are effective in imparting a satisfying (that is, rich taste) to the drink. .

Reference Example 2: Preparation of Peptide Fraction Providing Preferable Flavor to Beer-taste Beverage (1) Preparation of Peptide Fraction A commercially available beer-taste beverage was degassed, weighed, and freeze-dried. The lyophilized product was dissolved in a 100 mM NaCl solution to prepare a 5-fold concentrated solution, which was used as a sample for gel filtration fractionation. Gel filtration fractionation was carried out as described in Reference Example 1(2), and the resulting fraction C (fraction numbers: F25-29) was purified as described in Reference Example 1(3). At this time, the fraction that passed through the solid-phase extraction column was also recovered using a synthetic adsorbent (Separbeads SP207, manufactured by Mitsubishi Chemical Co., Ltd.) in order to improve the recovery rate in the purification. Protein quantification was performed by the Lowry method described in Reference Example 1 (4).

(2) Preparation of gel filtration fraction for HPLC analysis Dissolve the lyophilized product of (1) above in 50 mM sodium phosphate buffer (pH 7.0, containing 150 mM NaCl) to prepare a 2.5-fold concentrated solution, A sample for gel filtration fractionation was used. The peptide fraction was dissolved in the above buffer solution to a concentration equivalent to 10-fold concentration and used as a sample for gel filtration fractionation. Each of the fractionated samples was subjected to gel filtration under the following conditions to obtain fractions (fractions 1 to 10).

The conditions for the gel filtration method for HPLC analysis were as follows.
<Analysis conditions for gel filtration method for HPLC analysis>
Column: Superdex 75 10/300 (manufactured by GE Healthcare)
Sample injection volume: 100 μL
Eluent composition: 50 mM sodium phosphate (pH 7.0), 20% (v/v) acetonitrile, 150 mM NaCl
Flow rate: 0.5 mL/min (constant flow rate)
Detection wavelength: 215 nm

Protein quantification of the fraction was performed by the Lowry method described in Reference Example 1(4).

(3) Measurement of Molecular Weight by Gel Filtration for HPLC Analysis a. Sample Preparation for Gel Filtration for HPLC Analysis The fractionated fractions (fractions 1 to 10) obtained in (2) above were used as samples.

B. Creation of a molecular weight calibration curve Under the column, eluent, flow rate, and detection wavelength described in the gel filtration conditions for HPLC analysis, 50 μL of a peptide with a known molecular weight dissolved at 0.1 to 5 mg/mL in ultrapure water is injected. HPLC analysis was performed to confirm the retention time (Table 2). A calibration curve (Fig. 1) was prepared from the retention time and molecular weight.

C. Calculation of molecular weight As a result of molecular weight measurement, it was confirmed that the peptide contained in fraction 7 among fractions 1 to 10 had a molecular weight in the range of 800 to 1500 Da (gel filtration method for HPLC analysis).

(4) Calculation of ratio of peptides with molecular weights of 800 to 1500 Da Mass of peptides with molecular weights of 800 to 1500 Da (gel filtration method for HPLC analysis) in beverages relative to mass of total peptides derived from beverages fractionated by gel filtration method for HPLC analysis The ratio (percentage) of (peptide ratio) was determined by the following formula using the peptide concentration corresponding to the product in each fraction.

Example 1: Analysis of the effects of peptides and non-utilizable sugars on the flavor of sugar-free beer-taste fermented alcoholic beverages (1)
(1) Sugar-free beer-taste fermented alcoholic beverage The sugar-free beer-taste fermented alcoholic beverage of this example is a commercially available beer-taste beverage (malt usage ratio less than 25%, raw materials used: malt, hops, barley, sugars etc., sugar 0.2 g/100 mL) was used as the base beverage.

(2) Peptides with a molecular weight of 800 to 1500 Da Fraction 7 peptides obtained in Reference Example 2 were used as peptides with a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis).

(3) Non-utilizable sugars Isomaltose (97.0% purity) and panose (99.4% purity) were used as non-utilizable sugars.

(4) Component Analysis Component analysis of peptides with a molecular weight of 800 to 1500 Da (HPLC analysis gel filtration method) in the beverage was performed according to Reference Example 2. The concentration of peptides with a molecular weight of 800-1500 Da (gel filtration method for HPLC analysis) in the beverage was determined by the Lowry method. The concentrations of isomaltose and panose in beverages were determined by ion chromatography. Specifically, various samples were diluted with ultrapure water to a predetermined ratio and filtered through a 0.45 μm filter to obtain a filtrate, which was used as a sample solution for analysis. The sample was analyzed using an ion chromatograph ICS-6000 (manufactured by Dionex). Concentrations of isomaltose and panose were calculated from calibration curves prepared using standards.

(5) Sensory evaluation A sample with an alcohol concentration of 4 v / v% by diluting the beer-taste fermented alcoholic beverage with zero sugar in (1) above and adding alcohol for alcoholic beverages (Daiichi Alcohol Co., Ltd., alcohol concentration 95%). A beverage (Sample No. 1) was prepared. The original extract concentration (OE concentration) of this sample beverage was 6.37°P. The original extract concentration (OE concentration) was measured according to the BCOJ beer analysis method (Beer Brewers Association (2013), 8.3.6, 8.4.3 and 8.5. The measurement was an alcoholizer (Anton (manufactured by Pearl Co., Ltd.).

To the sample beverage of Sample No. 1, the above (2) peptide and the above (3) non-utilizable sugar were added to the concentration shown in Table 3, taking into consideration the concentrations originally contained in the beverage. Various sample beverages were prepared (sample numbers 2-17). Sugar concentrations in these sample beverages were less than 0.5 g/100 mL.

Sensory evaluation was carried out by 5 trained panelists. Specifically, the "thickness of taste" and "off-taste" were evaluated on a 17-point scale from 1 to 9 in 0.5-point increments, and the average score of the five panelists was calculated. Here, the term "thickness of taste" refers to the flavor sensation recognized by the breadth of taste, the complexity of the taste, and the imparting of a body feeling. As for "thickness of taste", the commercially available product of sample number 1 was given 1 point, and a score of 1.5 or more was judged as "slightly thick taste". "Miscellaneous taste" was evaluated by comprehensively evaluating astringent taste, harsh taste, and astringent taste. As for "off-taste", the commercial product of sample number 1 was given 3 points, and it was judged that the off-taste decreased as the score decreased.

(6) Results The concentrations of various components in the beverage and the results of sensory evaluation were as shown in Table 3 and FIG.

From the results in Table 3, the "thickness of taste" imparted by peptides with a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis) increased concentration-dependently by adding non-utilizable sugars (isomaltose and panose). It was confirmed that the "unpleasant taste" caused by the peptide was reduced in a concentration-dependent manner as well as further improved. However, since adding 0.15 g/100 mL of non-utilizable sugar left a slight sweetness on the tongue, the upper limit of the concentration of non-utilizable sugar in the beverage should be 0.15 g/100 mL. considered desirable. That is, from the results in Table 3, the concentration of peptides with a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis) in beer-taste fermented alcoholic beverages was adjusted to a range of 0.136 mg / mL or more, and isomaltose and It was shown that by adjusting the total concentration of panose in the range of 0.01 to 0.15 g/100 mL, it is possible to impart a rich taste while suppressing the off-taste caused by the peptides.

Example 2: Production of a sugar-free beer-taste fermented alcoholic beverage and analysis of the effects of peptides and non-utilizable sugars in the beverage on the flavor (1) Production of a sugar-free beer-taste fermented alcoholic beverage This example manufactured a beer-taste fermented alcoholic beverage. Barley malt was used as the main raw material for production (50% malt content). For saccharification, an enzyme preparation mainly composed of glucoamylase was used, and the temperature and time of saccharification were adjusted so that changes in the concentrations of isomaltose and panose reached equilibrium, followed by filtration to obtain wort.

Following the above wort adjustment step, liquid sugar mainly composed of hops and assimilable sugar is added to the obtained wort, boiled at 100 ° C., the wort is allowed to stand, and the trube is separated. After that, it was cooled to obtain a pre-fermentation liquid. After that, bottom-fermenting yeast was added to the pre-fermentation liquid, and main fermentation and post-fermentation were carried out according to a conventional method. Subsequently, the fermented liquor of the post-fermentation was stored by maintaining it at a lower temperature and filtered to obtain a clear beer-taste fermented alcoholic beverage.

(2) Component Analysis Concentration measurements of various components in the beer-taste fermented alcoholic beverage obtained in (1) above were performed according to the procedures described in Reference Example 2 and Example 1.

(3) Sensory Evaluation Sensory evaluation of the beer-taste fermented alcoholic beverage obtained in (1) above was performed according to the procedure described in Example 1 (5).

(4) Results The concentration of peptides with a molecular weight of 800 to 1500 Da (HPLC analysis gel filtration method) in the beer-taste fermented alcoholic beverage obtained in (1) above was 0.37 mg / mL, and the total of isomaltose and panose The concentration was 0.03 g/100 mL and the OE concentration was 6.6°P.

The beer-taste fermented alcoholic beverage obtained in the above (1) had an evaluation score of 4.9 for "thickness of taste" and an evaluation score of 2.4 for "off-taste". That is, the results of this example also show that the concentration of peptides with a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis) in beer-taste fermented alcoholic beverages is adjusted to a range of 0.136 mg / mL or more, and iso It was shown that by adjusting the total concentration of maltose and panose within the range of 0.01 to 0.15 g/100 mL, it is possible to impart a rich taste while suppressing the off-taste caused by the peptides.

A beer-taste fermented alcoholic beverage was separately prepared according to the above (1), and component analysis was performed according to the procedures of Reference Example 2 and Example 1. The concentration of peptides with a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis) is 0.37 mg/mL, and the ratio of the mass of peptides with a molecular weight of 800 to 1500 Da (HPLC analytical gel filtration method) in the beverage to the mass of all beverage-derived peptides fractionated by the HPLC analytical gel filtration method is 17.3%, the total concentration of isomaltose and panose was 0.03 g/100 mL, and the alcohol concentration was 3.72 v/v %.

Claims (5)

A beer-taste fermented alcoholic beverage containing at least a portion of malt as a raw material and having a carbohydrate concentration of less than 0.5 g/100 mL, wherein the peptide concentration in the beverage has a molecular weight of 800 to 1500 Da (HPLC analysis gel filtration method). is 0.136 mg / mL or more, the peptide is derived from a beer-taste fermented alcoholic beverage at least partly made from malt , and the total concentration of isomaltose and panose is 0.02 to 0.15 g / A beer-taste fermented alcoholic beverage of 100 mL. The beer-taste fermented alcoholic beverage according to claim 1, wherein the malt content is 50% or more. A method for producing a beer-taste fermented alcoholic beverage having an improved flavor and a sugar concentration of less than 0.5 g/100 mL, using malt as at least a portion of the raw material, wherein the beverage has a molecular weight of 800 to 1500 Da (HPLC Analytical gel filtration method) is adjusted to a peptide concentration of 0.136 mg / mL or more, and the peptide is derived from a beer-taste fermented alcoholic beverage that uses malt as at least a part of the raw material, and is the sum of isomaltose and panose A manufacturing method comprising adjusting the concentration to 0.02-0.15 g/100 mL. In the process of producing a beer-taste fermented alcoholic beverage, a step of adding a peptide with a molecular weight of 800 to 1500 Da (gel filtration method for HPLC analysis) contained in a beer-taste fermented alcoholic beverage containing malt as at least a part of the raw material and / or an iso 4. The production method according to claim 3, comprising adding maltose and/or panose. A method for improving the flavor of a beer-taste fermented alcoholic beverage with a carbohydrate concentration of less than 0.5 g/100 mL, wherein at least part of the raw material is malt , wherein the molecular weight of the beverage is 800 to 1500 Da (gel filtration method for HPLC analysis ) is adjusted to a peptide concentration of 0.136 mg / mL or more, the peptide is derived from a beer-taste fermented alcoholic beverage at least partly made from malt , and the total concentration of isomaltose and panose is 0.02 adjusting to ˜0.15 g/100 mL.

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