Method for producing beer-like carbonated beverage

Abstract

To provide a Lactobacillus breeding inhibitor which can inhibit breeding of Lactobacillus which is beer destructive bacteria in beer-like sparkling drink while suppressing influence for the quality of the drink, and the beer-like sparkling drink using the breeding inhibitor and a manufacturing method thereof.SOLUTION: A Lactobacillus breeding inhibitor for beer-like sparkling drink contains polylysine as an active component. The beer-like sparkling drink contains polylysine. A manufacturing method of the beer-like sparkling drink uses polylysine as a raw material.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to an antibacterial agent that inhibits the growth of lactic acid bacteria in a beer-like sparkling beverage, as well as a beer-like sparkling beverage using said antibacterial agent and a method for producing the same. [Background technology]

[0002] Beer is known to be a beverage in which microorganisms do not easily grow. This is due to its low pH, low dissolved oxygen content, alcohol content, and the use of hops. Hops impart a unique bitterness and aroma to beer, and their bitter components have antibacterial activity and suppress the growth of lactic acid bacteria, which cause beer to spoil.

[0003] Due to the diversification of consumer preferences, the ingredients and manufacturing methods used for beer-like sparkling beverages are becoming more diverse, and the development of beer-like sparkling beverages that do not use hops is also being considered. However, beer-like beverages that do not use hops may have a higher microbial risk than beer-like beverages that do use hops. Even when hops are used, there is a possibility that hop-resistant lactic acid bacteria may grow.

[0004] As a method for reducing the risk of microorganisms, in addition to environmental management, heat sterilization is considered. However, if harmful bacteria in beer grow during fermentation and storage, quality deterioration such as the generation of unpleasant odors and viscosity occurs, and heat sterilization in a later process may not be able to deal with this. In addition, heat sterilization may cause an undesirable flavor in beer-like sparkling beverages. For example, Patent Document 1 describes that in beer-flavored beverages that do not use hops as raw materials, the antibacterial effect of hops cannot be utilized, so sterilization of microorganisms may be required, for example, a heating process is performed, and that in beer-flavored beverages that do not use hops as raw materials, the unpleasant odor caused by the heating process is masked by including one or more selected from the group consisting of linalool, 2,5-dimethyl-4-hydroxy-3(2H)-furanone, and 2-acetylpyrazine.

[0005] On the other hand, polylysine is a homopolymer in which 25 to 35 ε-amino acids and α-carboxyl groups of L-lysine are linked by isopeptide bonds. Polylysine is known to have a growth inhibitory effect on various microorganisms such as gram-positive bacteria, gram-negative bacteria, and yeast (see Non-Patent Document 1), and is used as a food preservative. [Prior art documents] [Patent documents]

[0006] [Patent Document 1] JP 2018-023295 A [Non-patent literature]

[0007] [Non-Patent Document 1] Masamichi Muto, "Scientific Food Preservation Technology 5, Part 2) Existing Preservatives 1, ε-Polylysine," Antibacterial and Antifungal Agents Journal, Vol. 37, No. 10, 2009, pp. 765-772. Summary of the Invention [Problem to be solved by the invention]

[0008] Apart from environmental management and heat sterilization, a method for reducing the risk of microorganisms is to add an antimicrobial agent to beverages or products during the manufacturing process. However, simply adding an antimicrobial material to beer-like beverages produced through a fermentation process using yeast will not only inhibit the growth of lactic acid bacteria, which are harmful to beer, but also the growth of yeast, which is necessary for fermentation. In this case, poor fermentation will occur, causing problems.

[0009] The present invention aims to provide a lactic acid bacteria growth inhibitor that can inhibit the growth of lactic acid bacteria, which are harmful bacteria to beer, in a beer-like sparkling beverage while minimizing the impact on the quality of the beverage, as well as a beer-like sparkling beverage and a method for producing the same that use the antibacterial agent. [Means for solving the problem]

[0010] The present inventors have found that polylysine has a strong inhibitory effect on the proliferation of lactic acid bacteria in beer-like sparkling beverages, but a weak inhibitory effect on the proliferation of brewer's yeast, and have thus completed the present invention.

[0011] The present invention relates to R The method for producing a beer-like carbonated beverage includes the following steps [1] to [ 3 ]. [ 1 ] Polylysine is used as a raw material, It has a brewing process, a fermentation process, a storage process, and a filtration process. A method for producing a beer-like sparkling beverage, comprising adding the polylysine to a fermentation raw material liquid in the preparation step, adding the polylysine to a fermentation liquid in the fermentation step, or adding the polylysine to a fermentation liquid in the storage step. [ 2 ] The above [ 1 ] A method for producing a beer-like sparkling beverage. [ 3 The polylysine content of the produced beer-like carbonated beverage is 0.0001% (w / v) or more. [1] or [2] A method for producing a beer-like sparkling beverage. Effect of the Invention

[0012] The lactic acid bacteria proliferation inhibitor for beer-like sparkling beverages according to the present invention has a high inhibitory effect on the proliferation of lactic acid bacteria but a relatively weak inhibitory effect on the proliferation of brewer's yeast. Therefore, by using the lactic acid bacteria proliferation inhibitor, it is possible to reduce the risk of microorganisms while suppressing quality deterioration due to poor fermentation, not only in non-fermented beer-like sparkling beverages but also in fermented beer-like sparkling beverages. [Brief description of the drawings]

[0013] [Figure 1] 1 is a graph showing the results of measuring the absorbance at 600 nm over time for wort (A) to which L. brevis was added and for wort (B) to which S. pastorianus was added in Reference Example 1. [Diagram 2] 1 is a graph showing the results of measuring the absorbance at 600 nm over time for solutions in which L. brevis was added to each polylysine raw material solution in Example 1. [Diagram 3] 1 is a graph showing the results of measuring the absorbance at 600 nm over time for solutions in which S. pastorianus was added to each polylysine raw material solution in Example 1. [Figure 4] FIG. 1 shows the results of measuring the bacterial concentration of S. pastorianus and extract consumption during fermentation for a control sample (A) obtained by alcoholic fermentation of wort containing no polylysine, and a test sample (B) obtained by alcoholic fermentation of wort containing a final polylysine concentration of 0.0005% (w / v) in Example 3. [Diagram 5] 1 is a graph showing the results of measuring the absorbance at 600 nm over time for solutions in which two types of yeast were added to a polylysine solution having a final concentration of 0.0005% (w / v) in Example 4. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0014] In the present invention and the specification of this application, the term "beer-like sparkling beverage" refers to a beverage that has a beer-like taste and contains carbon dioxide. Furthermore, "beer-like" refers to a flavor that is reminiscent of beer, regardless of the product name or labeling. In other words, a beer-like sparkling beverage refers to a sparkling beverage that has a flavor, taste, and texture equivalent to or similar to that of beer, regardless of the alcohol content, the use of malt and hops, and the presence or absence of fermentation, and has high thirst quenching properties and drinkability.

[0015] The beer-like sparkling beverage according to the present invention may be an alcoholic beverage, or may be a so-called non-alcoholic beverage or low-alcohol beverage with an alcohol content of less than 1% by volume. It may also be a beverage made from malt, a beverage not made from malt, a beverage produced through a fermentation process, or a beverage produced without a fermentation process. Specific examples include beer, happoshu made from malt, sparkling alcoholic beverages not using malt, low-alcohol sparkling beverages, and non-alcoholic beer. In addition, it may be a liqueur or spirits obtained by mixing a beverage made from malt through a fermentation process with an alcohol-containing distillate. The alcohol-containing distillate is a solution containing alcohol obtained by a distillation operation, and may be, for example, raw material alcohol, and distilled liquors such as spirits, whiskey, brandy, vodka, rum, tequila, gin, and shochu can be used.

[0016] The lactic acid bacteria proliferation inhibitor for beer-like sparkling beverages according to the present invention (hereinafter, sometimes referred to as "the lactic acid bacteria proliferation inhibitor of the present invention") is characterized by containing polylysine as an active ingredient. Polylysine has high proliferation inhibitory activity against lactic acid bacteria. For this reason, polylysine is very useful as an antibacterial agent used in beer-like sparkling beverages in which lactic acid bacteria are the main harmful bacteria. Also, polylysine is known to exhibit proliferation inhibitory activity against both lactic acid bacteria and yeast, but its proliferation inhibitory activity against lactic acid bacteria is stronger. For this reason, even in a fermented beer-like sparkling beverage, by containing polylysine at a concentration that exhibits sufficient antibacterial activity against lactic acid bacteria but has no or weak proliferation inhibitory activity against yeast, it is possible to suppress contamination due to proliferation of lactic acid bacteria while suppressing poor fermentation and the resulting deterioration in quality.

[0017] The polylysine used in the present invention may be extracted from a natural product, may be produced by a transformed microorganism, or may be chemically synthesized. For example, polylysine is produced by Streptomyces bacteria through fermentation, and can be extracted from this microorganism. The polylysine used in the present invention can also be commercially available. The commercially available polylysine may be a purified product of polylysine, or may be an additive for food and drink containing an active ingredient other than polylysine.

[0018] By adding the lactic acid bacteria growth inhibitor of the present invention to a beer-like sparkling beverage, the growth of lactic acid bacteria in the beer-like sparkling beverage can be inhibited. Lactic acid bacteria are the main harmful bacteria in beer-like sparkling beverages, and if the growth of lactic acid bacteria can be sufficiently inhibited, the microbial risk can be significantly reduced. The amount of the lactic acid bacteria growth inhibitor added to the beer-like sparkling beverage may be sufficient to obtain the effect of inhibiting the growth of lactic acid bacteria in the beverage (lactic acid bacteria inhibiting effect). For example, the lactic acid bacteria growth inhibitor can obtain a sufficient lactic acid bacteria inhibiting effect while suppressing quality deterioration due to poor fermentation by making the content of polylysine in the beverage preferably 0.0001% (w / v) or more, more preferably 0.0001 to 0.001% (w / v), even more preferably 0.0002 to 0.0005% (w / v), and even more preferably 0.0002 to 0.0004% (w / v). The content of polylysine in beer-like carbonated beverages and the like can be measured by a general method used for quantifying specific proteins in beverages. For example, the method involves converting the α-amino group of polylysine in a sample into a dansyl derivative, hydrolyzing the resulting α-dansyllysine (α-DNS-Lys), and quantifying the resulting α-dansyllysine by high performance liquid chromatography (HPLC) or the like (Takashi Hishiyama et al., "Quantitative Analysis of ε-Polylysine in Foods by Precolumn Dansyl Derivatization High Performance Liquid Chromatography," Journal of the Japanese Society for Food Science and Technology, October 1996, Vol. 43, No. 10, pp. 1105-1109).

[0019] The lactic acid bacteria growth inhibitor of the present invention has a high lactic acid bacteria inhibition effect, and therefore the lactic acid bacteria growth inhibitor of the present invention can be particularly suitably used in beer-like sparkling beverages that do not use hops as a raw material, i.e., beer-like sparkling beverages that do not contain antibacterial components derived from hops. By adding the lactic acid bacteria growth inhibitor, it is possible to sufficiently reduce the microbial risk even in beer-like sparkling beverages that do not use hops as a raw material.

[0020] Polylysine also exhibits a high growth inhibitory effect against so-called hop-resistant lactic acid bacteria, and therefore, by using the lactic acid bacteria growth inhibitor of the present invention, a sufficient lactic acid bacteria inhibitory effect can be exhibited even in beer-like sparkling beverages made from hops, and microbial risk can be sufficiently reduced.

[0021] The beer-like sparkling beverage according to the present invention can be produced in the same manner as a general fermented beer-like sparkling beverage or a non-fermented beer-like sparkling beverage, except that polylysine is contained. Hereinafter, a method for producing a general fermented beer-like sparkling beverage and a non-fermented beer-like sparkling beverage will be described.

[0022] Fermented beer-like sparkling beverages produced through a fermentation process can generally be produced through the steps of brewing (preparation of fermented raw material liquid), fermentation, storage, and filtration.

[0023] In the preparation process (fermentation raw material liquid preparation process), a fermentation raw material liquid is prepared from one or more selected from the group consisting of grain raw materials and carbohydrate raw materials. Specifically, a mixture containing the fermentation raw materials and raw material water is heated, starch is saccharified to prepare a sugar liquid, and the obtained sugar liquid is boiled, and then at least a part of the solid content is removed to prepare the fermentation raw material liquid.

[0024] First, a mixture containing at least one of a grain raw material and a sugar raw material and raw material water is prepared and heated, and the starch of the grain raw material is saccharified to prepare a sugar liquid. As the raw material for the sugar liquid, only the grain raw material may be used, only the sugar raw material may be used, or both may be used in combination. Examples of the grain raw material include barley, wheat, and malt thereof, as well as rice, corn, beans such as soybeans, and potatoes. The grain raw material can be used as grain syrup, grain extract, and the like, but it is preferable to use it as a grain pulverized product obtained by pulverization. The pulverization of grains can be performed by a conventional method. The grain pulverized product may be a product that has been subjected to a conventional process before or after pulverization, such as malt pulverized product, corn starch, corn grits, and the like. In the present invention, the grain pulverized product used is preferably malt pulverized product. By using malt pulverized product, a fermented beer-like sparkling beverage with a more pronounced beer-like flavor can be produced. The ground malt may be barley, for example two-rowed barley, germinated in a conventional manner, dried, and then ground to a predetermined particle size. The grain raw material used in the present invention may be one type of grain raw material or a mixture of multiple types of grain raw materials. For example, ground malt may be used as the main raw material, and ground rice or corn may be used as the secondary raw material. Examples of carbohydrate raw materials include sugars such as liquid sugar.

[0025] The mixture may contain auxiliary materials other than the grain raw material and water. Examples of the auxiliary materials include hops, dietary fiber, yeast extract, fruit juice, bittering agents, coloring agents, herbs, flavoring agents, etc. In addition, saccharifying enzymes such as α-amylase, glucoamylase, pullulanase, and enzyme agents such as protease can be added as necessary.

[0026] The saccharification treatment is carried out using enzymes derived from the grain raw materials, etc., or enzymes added separately. The temperature and time during the saccharification treatment are appropriately adjusted taking into consideration the type of grain raw materials, etc. used, the ratio of the grain raw materials to the entire fermentation raw materials, the type of enzymes added and the amount of the mixture, the quality of the desired fermented beer-like sparkling beverage, etc. For example, the saccharification treatment can be carried out by a conventional method, such as by holding a mixture containing the grain raw materials, etc. at 35 to 70°C for 20 to 90 minutes.

[0027] The sugar liquid obtained after saccharification can be boiled to prepare a broth (a boiled product of the sugar liquid). It is preferable to filter the sugar liquid before the boiling treatment, and to boil the obtained filtrate. Alternatively, instead of the filtrate of the sugar liquid, a mixture of malt extract and warm water may be used and boiled. The boiling method and conditions can be appropriately determined.

[0028] By adding herbs and the like as appropriate before or during the boiling process, a fermented beer-like sparkling beverage having a desired flavor can be produced. For example, by adding hops before or during the boiling process and performing the boiling process in the presence of hops, the flavor and aroma components of the hops, particularly the bitter components, can be efficiently boiled out. The amount of hops to be added, the manner of addition (for example, adding in several portions), and the boiling conditions can be appropriately determined.

[0029] The broth obtained after the boiling process contains residues such as proteins generated by precipitation. Therefore, at least a part of the solid contents such as residues is removed from the broth. The residues may be removed by any solid-liquid separation process, but generally, a tank called a whirlpool is used to remove the precipitate. The temperature of the broth at this time may be 15°C or higher, and generally, the temperature is about 50 to 100°C. The broth (filtrate) after removing the residues is cooled to an appropriate fermentation temperature by a plate cooler or the like. The broth after removing the residues becomes the fermentation raw material liquid.

[0030] Next, in the fermentation step, yeast is inoculated into the cooled fermentation raw material liquid to carry out fermentation. The cooled fermentation raw material liquid may be subjected to the fermentation step as it is, or may be subjected to the fermentation step after being adjusted to a desired extract concentration. The yeast used for fermentation is not particularly limited, and may be appropriately selected from yeasts normally used in the production of alcoholic beverages. Although top-fermenting yeast or bottom-fermenting yeast may be used, bottom-fermenting yeast is preferred because it is easily applicable to large-scale brewing equipment.

[0031] Furthermore, in the storage step, the obtained fermented liquid is aged in a storage tank and stored under low-temperature conditions of about 0°C for stabilization, and then in the filtration step, the aged fermented liquid is filtered to remove yeast and proteins insoluble in the temperature range, thereby obtaining the desired fermented beer-like sparkling beverage. The filtration process may be any method capable of filtering out yeast, and examples of the filtration include diatomaceous earth filtration and filter filtration using a filter with an average pore size of about 0.4 to 1.0 μm.

[0032] Polylysine can be added as a raw material in any step of the manufacturing process of fermented beer-like sparkling beverage. For example, it may be added to the fermentation raw material liquid in the preparation step, to the fermentation liquid in the fermentation step, or to the fermentation liquid in the storage step. Since polylysine retains the lactic acid bacteria inhibitory effect even after heat treatment, it may be added before or during the boiling treatment in the preparation step.

[0033] The pH of the fermentation raw material liquid is adjusted to 5.0 to 6.0 to facilitate precipitation of heat-coagulable proteins and to a pH suitable for alcoholic fermentation. The pH after fermentation is 4.0 to 4.5. Polylysine can inhibit the proliferation of lactic acid bacteria at a pH range of 4.0 to 6.0, preferably 4.0 to 5.5.

[0034] Non-fermented beer-like sparkling beverages produced without a fermentation process can generally be produced by a method of mixing each ingredient (blending method). For example, specifically, they can be produced by a blending process of preparing a blend by mixing each ingredient, and a gas introduction process of adding carbon dioxide gas to the obtained blend.

[0035] First, in the blending step, the raw materials are mixed to prepare a blended liquid. In the blending step, it is preferable to prepare a blended liquid in which all raw materials except carbon dioxide gas are mixed. The order in which the raw materials are mixed is not particularly limited. All raw materials may be added to the raw material water at the same time, or the raw materials may be added sequentially, such as by adding the raw materials remaining after dissolving the raw materials added earlier. In addition, for example, solid raw materials (e.g., powder or granular) and alcohol may be mixed with the raw material water, or the solid raw materials may be prepared as aqueous solutions in advance, and the aqueous solutions, alcohol, and raw material water, if necessary, may be mixed.

[0036] Examples of raw materials include bittering agents, acidulants, sweetening agents, caramel coloring, flavoring agents, ethanol (raw alcohol), emulsifiers, polysaccharides, water-soluble dietary fiber, proteins or their hydrolyzates, etc.

[0037] Examples of the acidulant include lactic acid, citric acid, gluconic acid, tartaric acid, malic acid, succinic acid, phosphoric acid, adipic acid, and fumaric acid. These acidulants may be used alone or in combination of two or more.

[0038] Examples of bittering agents include hops, iso-α acid, tetraiso-α acid, oxides of β acid, magnesium salts, calcium salts, naringin, quasin, quinine, momordicine, quercitrin, theobromine, caffeine, bitter melon, Swertia japonica tea, Kuding tea, Artemisia absinthium extract, gentiana extract, cinchona extract, etc. These bittering agents may be used alone or in combination of two or more.

[0039] Hops used as a raw material may be fresh hops, dried hops, hop pellets, or a hop processed product. The hop processed product may be a hop extract obtained by extracting bitter components from hops, or may be an isoform hop extract, or a hop processed product containing components obtained by isoforming bitter components in hops, such as tetrahydroisohumulone and hexahydroisohumulone.

[0040] Examples of sweeteners include sucrose, glucose, fructose, isomerized liquid sugar, and high-intensity sweeteners. Examples of high-intensity sweeteners include aspartame, sucralose, acesulfame potassium, neotame, stevia, and enzyme-treated stevia. These sweeteners may be used alone or in combination of two or more.

[0041] Examples of the flavoring include beer extract, beer flavoring, hop flavoring, etc. These flavorings may be used alone or in combination of two or more kinds.

[0042] Examples of emulsifiers include polyglycerin fatty acid esters, glycerin fatty acid esters, sucrose fatty acid esters, polypropylene glycol fatty acid esters, sorbitan fatty acid esters, polysorbates, and the like.

[0043] Examples of polysaccharides include starch and dextrin. Dextrin is a carbohydrate obtained by hydrolyzing starch and is larger than oligosaccharides (carbohydrates formed by polymerizing about 3 to 10 monosaccharides). These polysaccharides may be used alone or in combination of two or more kinds.

[0044] Water-soluble dietary fiber means carbohydrates that dissolve in water and are not or are difficult to digest by human digestive enzymes. Examples of water-soluble dietary fiber include soybean dietary fiber (soluble soybean polysaccharides), polydextrose, indigestible dextrin, galactomannan, inulin, guar gum hydrolyzate, pectin, gum arabic, etc. These water-soluble dietary fibers may be used alone or in combination of two or more types.

[0045] When insoluble matter is generated in the preparation liquid prepared in the preparation step, it is preferable to subject the preparation liquid to a treatment for removing the insoluble matter, such as filtration, before the gas introduction step. The insoluble matter removal treatment is not particularly limited, and can be performed by a method commonly used in the technical field, such as a filtration method or a centrifugation method. In the present invention, it is preferable to remove the insoluble matter by filtration, and it is more preferable to remove the insoluble matter by diatomaceous earth filtration.

[0046] Next, in the gas introduction step, carbon dioxide gas is added to the preparation obtained in the blending step. This results in a non-fermented beer-like sparkling beverage. The addition of carbon dioxide imparts a refreshing feeling similar to that of beer. The addition of carbon dioxide gas can be carried out by a conventional method. For example, the preparation obtained in the blending step and carbonated water may be mixed, or carbon dioxide gas may be directly added to and dissolved in the preparation obtained in the blending step.

[0047] After the addition of carbon dioxide gas, the resulting non-fermented beer-like sparkling beverage may be subjected to a further treatment such as filtration to remove insoluble matter. The insoluble matter removal treatment is not particularly limited, and may be carried out by a method commonly used in the art.

[0048] Polylysine can be added as a raw material in any step of the production process of the non-fermented beer-like sparkling beverage. For example, it may be added to the preparation liquid together with other raw materials in the preparation step, or it may be added after the introduction of carbon dioxide gas, before or after the insoluble matter removal treatment.

[0049] When the sparkling beverage according to the present invention is a packaged beverage, the container in which the sparkling beverage according to the present invention is filled is not particularly limited. Specific examples include glass bottles, cans, and flexible containers. Flexible containers include containers made by molding flexible resins such as PE (polyethylene), PP (polypropylene), EVOH (ethylene-vinyl alcohol copolymer), and PET (polyethylene terephthalate). Flexible containers may be made of a single-layer resin or a multi-layer resin. EXAMPLES

[0050] The present invention will now be described in more detail with reference to examples and reference examples, but the present invention is not limited to the following examples.

[0051] [Reference example 1] The growth ability of brewer's yeast (Saccharomyces pastorianus: hereafter referred to as "S. pastorianus") and an environmentally isolated strain of lactic acid bacteria (Lactobacillus brevis: hereafter referred to as "L. brevis") was investigated in wort containing hops and in wort prepared without hops.

[0052] <Microbial pre-culture and bacterial solution preparation> S. pastorianus was inoculated onto CP-added potato dextrose agar medium (Eiken Chemical Co., Ltd.), and L. brevis was inoculated onto MRS liquid medium (de Man, Rogosa and Sharpe, Merck), and each was cultured at 25°C for 3 days. The pre-cultured bacteria was cultured at a concentration of approximately 10 5 The bacteria were suspended in sterile saline so that each cell was divided into individual cells, and the suspension was used as the bacterial solution.

[0053] <Preparation of wort> A mixture of 40 kg of malt and 160 L of water was kept at 58°C for 20 minutes to decompose proteins, and then kept at 64°C for 20 minutes to saccharify the malt-derived components. The mixture after saccharification was filtered to obtain clear wort that did not contain hops.

[0054] The obtained wort was heat sterilized in an autoclave (121°C, 15 minutes), and the trub was removed using filter paper. The wort after trub removal was adjusted to pH 5.25±0.05 using 5N HCl or NaOH, and then sterilized using a filter (pore size 0.45 μm) to obtain hop-free wort. The same procedure was followed except that hops were added during heat sterilization in an autoclave, and the resulting hop-containing wort was sterilized by filtration.

[0055] <Microbial culture> Hop-free or hop-containing wort was inoculated with S. pastorianus or L. brevis at 10 per well in a 96-well plate. 3 The plates were inoculated at 100 μL / cell and cultured at 25° C., and the absorbance at 600 nm was measured over time. The absorbance was measured using a VICTOR Nivo Multimode Plate Reader (manufacturer: PerkinElmer, S / N: HH35L3517059, excitation filter: 600 / 10 nm). A sample not inoculated with the microorganism was used to measure the background.

[0056] The measurement results for the wort to which L. brevis was added are shown in Figure 1(A), and the measurement results for the wort to which S. pastorianus was added are shown in Figure 1(B). As a result, growth of S. pastorianus was observed in both worts, while growth of L. brevis was confirmed in the unhopped wort, as indicated by an increase in absorbance, but no growth was observed in the hopped wort.

[0057] [Example 1] The proliferation ability of S. pastorianus and L. brevis used in Reference Example 1 in the presence of polylysine was examined.

[0058] <Preparation of polylysine raw material> Polylysine was prepared by diluting an aqueous solution (10% (w / v) polylysine solution) prepared with pure water to a final concentration of 10% (w / v) with hop-free wort to a final polylysine concentration of 0.0010% (w / v), adjusting the pH to 5.25±0.05 with hydrochloric acid or lactic acid, and then sterilizing the solution with a filter (pore size 0.45 μm). The obtained 0.0010% (w / v) polylysine solution was further diluted with hop-free wort to prepare polylysine solutions with final polylysine concentrations of 0.00001% (w / v), 0.00002% (w / v), 0.00004% (w / v), 0.00006% (w / v), and 0.00008% (w / v). The sample used for evaluating heat resistance was prepared in the same manner as above using a 10% (w / v) polylysine solution heated at 100° C. for 30 minutes.

[0059] <Measurement of antibacterial performance> The bacterial solution prepared in the same manner as in Reference Example 1 was inoculated into a 96-well plate (8 rows × 12 columns) at 10 μL per well, and 90 μL of the polylysine raw material solution adjusted to each concentration was added, followed by culturing under anaerobic conditions for 7 days at 25° C. The viable cell count (test cell count) of the inoculum was measured by a plate spread culture method using CP-added potato dextrose agar medium or MRS agar medium, and calculated by converting it into the viable cell count of the inoculum. The absorbance at 600 nm of the solution in each well was measured over time in the same manner as in Reference Example 1 to examine the proliferation of microorganisms.

[0060] Among the samples seeded with L. brevis, the results of the polylysine solution prepared from the unheated 10% (w / v) polylysine solution are shown in Figure 2(A), and the results of the polylysine solution prepared from the heated 10% (w / v) polylysine solution are shown in Figure 2(B). Among the samples seeded with S. pastorianus, the results of the polylysine solution prepared from the unheated 10% (w / v) polylysine solution are shown in Figure 3(A), and the results of the polylysine solution prepared from the heated 10% (w / v) polylysine solution are shown in Figure 3(B).

[0061] As a result, it was confirmed that the increase in absorbance was suppressed in all samples inoculated with L. brevis compared to the results for the hop-free wort in Reference Example 1, and the proliferation of L. brevis was inhibited. Furthermore, an increase in absorbance was confirmed in all samples inoculated with S. pastorianus, confirming that the proliferation of S. pastorianus is possible.

[0062] In particular, in a solution containing 0.0004% (w / v) polylysine, the proliferation of L. brevis was hardly observed regardless of whether the polylysine was unheated or heat-treated (FIGS. 2(A) and (B)), whereas S. pastorianus showed sufficient proliferation (FIGS. 3(A) and (B)). From these results, it was confirmed that the proliferation of lactic acid bacteria can be suppressed while suppressing the effect on fermentation by adding an appropriate amount of polylysine.

[0063] [Example 2] The effect of polylysine on alcohol fermentation by yeast was investigated.

[0064] <Preparation of polylysine raw material> A commercially available polylysine preparation for food additives (product name: "Sunkeeper No. 381", manufactured by San-Ei Gen Co., Ltd., composition: polylysine 50% (w / w), glycerin 50% (w / w)) was used to prepare the polylysine raw material. The polylysine preparation was added to hop-free wort so that the final polylysine concentration was 0.0005, 0.00125, 0.0025, or 0.005% (w / v), and this was used as the polylysine raw material.

[0065] <Inoculation of test bacteria> The number of S. pastorianus cells precultured in the same manner as in Example 1 was approximately 40 × 10 8 The resulting bacterial solution was inoculated in an Erlenmeyer flask in an amount of 10 mL, and 190 mL of the polylysine raw material adjusted to each concentration was added and mixed, and the flask was plugged and cultured at 10.5° C. for 8 days under aerobic conditions.

[0066] Alcoholic fermentation produces two molecules of ethanol and two molecules of carbon dioxide from one molecule of sugar [C6H 12 O6 (180g) → 2C2H5OH (46g x 2) + 2CO2 (44g x 2)]. Therefore, the concentration (g / L) of alcohol produced by alcoholic fermentation can be expressed by the following formula (1). Furthermore, since the specific gravity of ethanol is 0.789g / mL, it can be converted to volumetric concentration as shown in the following formula (2). Therefore, the amount of carbon dioxide reduction was obtained by measuring the weight of the Erlenmeyer flask during cultivation, and the concentration of alcohol produced (% (v / v)) was estimated.

[0067] Formula (1): [Alcohol concentration of fermentation liquid (g / L)] = ((46 / 44) × [amount of carbon dioxide reduced (g)]) / [volume of culture liquid (L)] Equation (2): [Alcohol concentration of fermentation liquid (% (v / v))] = (0.1325 × [amount of carbon dioxide reduced (g)]) / [volume of culture liquid (L)]

[0068] The measurement results of the alcohol concentration (% (v / v)) of each reaction solution are shown in Table 1. The amount of alcohol produced decreased as the final concentration of polylysine in the fermentation solution increased. However, even when the final concentration of polylysine was 0.005%, the alcohol concentration of the fermentation solution was about 0.7% (v / v), and it was confirmed that alcohol was produced without any problems.

[0069] [Table 1]

[0070] [Example 3] Alcoholic fermentation was carried out by mass culture in the presence of polylysine, and the resulting fermented beer-like sparkling beverage was subjected to a sensory evaluation. The S. pastorianus and L. brevis used were those used in Reference Example 1.

[0071] <Production of fermented beer-like sparkling beverage> 22 kg of cornstarch and 66 kg of water were mixed and heated to swell (swollen starch). Next, a mixture of 44 kg of malt and 110 kg of water was kept at 55°C for 30 minutes to carry out protein decomposition treatment. The swollen starch was then added to the mixture, and the mixture was kept at 64.5°C for 50 minutes to saccharify the malt and cornstarch-derived components, and filtered to obtain clear wort. Thereafter, the wort was boiled for 80 minutes without adding hops, and prepared hot water was added. The polylysine preparation used in Example 2 was added to the resulting boiled wort so that the final concentration of polylysine was 0.0005% (w / v), and the mixture was stirred, and the thermal coagulum was removed and then cooled to about 8°C. The cooled hop-free wort was transferred to a fermentation tank, and the bacterial counts of S. pastorianus and L. brevis were 20 x 10 per mL, respectively. 6 pcs and 1.0×10 5 The cells were added so that each cell was equal to one cell, and the cells were cultured at 10.5°C.

[0072] The bacterial concentration and extract consumption of S. pastorianus during fermentation were measured. The measurement results are shown in FIG. 4. FIG. 4(A) shows the result of a control sample in which wort without added polylysine was subjected to alcoholic fermentation, and FIG. 4(B) shows the result of a test sample in which wort with a final polylysine concentration of 0.0005% (w / v) was subjected to alcoholic fermentation. In addition, sampling was performed every 7 days of fermentation to observe the viable cell count of L. brevis. The measurement results are shown in Table 2.

[0073] [Table 2]

[0074] As shown in Figure 4, in wort containing polylysine at a final concentration of 0.0005% (w / v), S. pastorianus grew and consumed the extract in the same way as in the control. Moreover, as shown in Table 2, the viable cell mass of L. brevis gradually decreased during the fermentation and maturation period of wort containing polylysine at a final concentration of 0.0005% (w / v).

[0075] <Sensory evaluation> The obtained test sample and control sample were first clarified by filtration, diluted 1.2 times, and then filled into a container. Next, to examine whether the addition of polylysine caused any change in flavor, a sensory evaluation was performed on both samples using a three-point test method. The sensory evaluation was performed by a panel of 11 experts according to the method described in "BCOJ Sensory Evaluation Method 2002." The evaluation results are shown in Table 3.

[0076] [Table 3]

[0077] As shown in Table 3, of the 11 panelists, only two panelists correctly distinguished the test sample from the control sample, and no significant difference was observed between the two samples. In other words, even when alcoholic fermentation was performed in the presence of polylysine, a fermented beer-like sparkling beverage with excellent flavor could be produced, just like in the absence of polylysine.

[0078] [Example 4] The growth ability of two types of yeast other than S. pastorianus in the presence of polylysine was examined. The yeasts used were Saccharomyces cerevisiae (hereinafter referred to as "S. cerevisiae") and Saccharomyces bayanus (hereinafter referred to as "S. bayanus").

[0079] The yeast solution was placed in a well of a 96-well plate together with polylysine at a final concentration of 0.0005% (w / v), and cultured under anaerobic conditions at 25° C. for 7 days in the same manner as in Example 1, except that S. cerevisiae or S. bayanus was used instead of S. pastorianus as the yeast, and the polylysine raw material solution prepared in Example 1 or the commercially available food additive polylysine preparation (product name: "Sunkeeper No. 381", manufactured by San-Ei Gen Co., Ltd.) used in Example 2 was used as the polylysine raw material. The yeast solution was placed in a well of a 96-well plate together with polylysine at a final concentration of 0.0005% (w / v), and cultured under anaerobic conditions at 25° C. for 7 days in the same manner as in Example 1. The absorbance at 600 nm of each well was measured over time to examine the growth of the microorganisms.

[0080] The results of measuring the absorbance at 600 nm for each well are shown in Figure 5. The changes in absorbance of S. cerevisiae with the addition of a polylysine preparation (in the figure, "S. cerevisiae / polylysine preparation") and S. cerevisiae with the addition of a polylysine raw material solution (in the figure, "S. cerevisiae / polylysine raw material solution") were almost the same, and S. cerevisiae showed sufficient growth in the presence of 0.0005% (w / v) polylysine. Similarly, the changes in absorbance of S. bayanus with the addition of a polylysine preparation (in the figure, "S. bayanus / polylysine preparation") and S. bayanus with the addition of a polylysine raw material solution (in the figure, "S. bayanus / polylysine raw material solution") were almost the same, and S. bayanus also showed sufficient growth in the presence of 0.0005% (w / v) polylysine.

Claims

1. Using polylysine as a raw material, It has a brewing process, a fermentation process, a storage process, and a filtration process. A method for producing a beer-like sparkling beverage, comprising adding the polylysine to a fermentation raw material liquid in the preparation step, adding the polylysine to a fermentation liquid in the fermentation step, or adding the polylysine to a fermentation liquid in the storage step.

2. The method for producing a beer-like sparkling beverage according to claim 1, which does not use hops as a raw material.

3. 3. The method for producing a beer-like sparkling beverage according to claim 1 or 2, wherein the polylysine content in the produced beer-like sparkling beverage is 0.0001% (w / v) or more.

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