JP2020156462A - Lactobacillus breeding inhibitor for beer-like sparkling drink - Google Patents

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

The present invention relates to an antibacterial agent that suppresses the growth of lactic acid bacteria in a beer-like effervescent beverage, a beer-like effervescent beverage using the antibacterial agent, and a method for producing the same.

Beer is known to be a beverage in which microorganisms do not easily grow. This is due to the low pH of beer, low dissolved oxygen, alcohol content, and the use of hops. Hops impart the bitterness and aroma peculiar to beer, and the bitterness component exhibits antibacterial activity and has a function of suppressing the growth of lactic acid bacteria that deteriorate beer.

Due to the diversification of consumer tastes, the materials and manufacturing methods used for beer-like sparkling beverages are also diversifying, and the development of beer-like sparkling beverages that do not use hops is also being considered. However, hop-free beer-like beverages may have an increased microbial risk compared to hop-based beer-like beverages. Even when hops are used, hop-resistant lactic acid bacteria may grow.

In addition to environmental management, heat sterilization can be considered as a method for reducing the risk of microorganisms. However, if harmful bacteria of beer grow during fermentation and storage, quality deterioration such as generation of unpleasant odor and thickening may occur, and heat sterilization in the subsequent process may not be possible. In addition, heat sterilization may give an unfavorable flavor to beer-like sparkling beverages. For example, in Patent Document 1, beer-taste beverages that do not use hops as raw materials may require sterilization of microorganisms because the antibacterial action of hops cannot be utilized. For example, a heating step is performed, and hops are used as raw materials. In a beer-taste beverage that does not use hops, by containing one or more selected from the group consisting of linalol, 2,5-dimethyl-4-hydroxy-3 (2H) -furanone and 2-acetylpyrazine, it is unpleasant due to the heating process. It is stated that the scent is masked.

On the other hand, polylysine is a homopolymer in which 25 to 35 ε-amino acids of L-lysine and α-carboxyl groups 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.

Japanese Unexamined Patent Publication No. 2018-023295

Masamichi Muto, "Scientific Preservative Technology for Foods 5 Details 2) Existing Preservatives 1 ε-Polylysine", Antibacterial and Antifungal Magazine, Vol. 37, No. 10, published in 2009, pp. 765-772.

As a method for reducing the risk of microorganisms other than environmental management and heat sterilization treatment, a method of adding an antibacterial agent against the microorganisms to a beverage or a product in the middle of a manufacturing process can be considered. However, in a beer-like beverage produced through a fermentation process using yeast, simply adding an antibacterial material suppresses the growth of not only lactic acid bacteria, which are harmful bacteria of beer, but also yeast necessary for fermentation. In this case, adverse effects due to poor fermentation occur.

The present invention is an agent for suppressing the growth of lactic acid bacteria, which is a harmful bacterium of beer, and a beer-like effervescent agent using the antibacterial agent, while suppressing the influence on the quality of the beer-like effervescent beverage. It is an object of the present invention to provide a beverage and a method for producing the same.

The present inventors have found that polylysine has a strong inhibitory effect on the growth of lactic acid bacteria in a beer-like effervescent beverage, but a weak inhibitory effect on the growth of brewer's yeast, and completed the present invention.

The methods for producing a lactic acid bacterium growth inhibitor, a beer-like effervescent beverage, and a beer-like effervescent beverage according to the present invention are as follows [1] to [8].
[1] A lactic acid bacterium growth inhibitor for beer-like effervescent beverages, which comprises polylysine as an active ingredient.
[2] A beer-like effervescent beverage characterized by containing polylysine.
[3] The beer-like effervescent beverage according to [2] above, wherein the content of polylysine is 0.0001% (w / v) or more.
[4] The beer-like sparkling beverage according to the above [2] or [3], which does not use hops as a raw material.
[5] The beer-like effervescent beverage according to any one of [2] to [4] above, which is a fermented beer-like effervescent beverage.
[6] A method for producing a beer-like effervescent beverage, which comprises using polylysine as a raw material.
[7] The method for producing a beer-like sparkling beverage according to the above [6], which does not use hops as a raw material.
[8] The method for producing a beer-like effervescent beverage according to the above [6] or [7], wherein the produced beer-like effervescent beverage has a polylysine content of 0.0001% (w / v) or more.

The lactic acid bacterium growth inhibitor for beer-like effervescent beverages according to the present invention has a high inhibitory effect on the growth of lactic acid bacteria, but has a relatively weak inhibitory effect on the growth of brewer's yeast. Therefore, by using the lactic acid bacterium growth inhibitor, not only non-fermented beer-like effervescent beverages but also fermented beer-like effervescent beverages can reduce the risk of microorganisms while suppressing quality deterioration due to poor fermentation and the like. be able to.

In Reference Example 1, it is a graph which showed the result of having measured the absorbance at 600 nm of the wort (A) to which L. brevis was added and the wort (B) to which S. pastorianus was added over time. It is a graph which showed the result of having measured the absorbance at 600 nm of the solution which added L. brevis to each polylysine raw material solution with time in Example 1. FIG. In Example 1, it is a graph which showed the result of having measured the absorbance at 600 nm of the solution which added S. pastorianus to each polylysine raw material solution with time. In Example 3, the control sample (A) in which the wort without polylysine was alcohol-fermented and the test sample (B) in which the wort having a final concentration of 0.0005% (w / v) of polylysine was alcohol-fermented. It is a figure which showed the result of having measured the bacterial concentration and extract consumption of S. pastorianus during fermentation. FIG. 5 is a graph showing the results of measuring the absorbance at 600 nm of a solution obtained by adding two types of yeast to a solution having a final concentration of polylysine of 0.0005% (w / v) in Example 4 over time.

In the present invention and the specification of the present application, the "beer-like effervescent beverage" means a beverage containing carbon dioxide gas, which has a beer-like character. In addition, "beer-likeness" means a taste reminiscent of beer in terms of flavor, regardless of the product name or label. That is, a beer-like effervescent beverage has a flavor, taste and texture equivalent to or similar to beer, regardless of the alcohol content, the use of malt and hops, and the presence or absence of fermentation. However, it means a beverage with a high sense of thirst and drinkability.

The beer-like effervescent beverage according to the present invention may be an alcoholic beverage, or may be a so-called non-alcoholic beverage or a low-alcoholic beverage having an alcohol content of less than 1% by volume. Further, it may be a beverage made from malt, a beverage not made from malt, a beverage produced through a fermentation step, or a beverage produced without undergoing a fermentation step. It may be. Specific examples thereof include beer, sparkling wine made from malt, sparkling alcoholic beverages that do not use malt, low alcohol sparkling beverages, and non-alcoholic beer. In addition, liqueurs or spirits obtained by mixing a beverage produced from malt as a raw material through a fermentation step with an alcohol-containing distillate may be used. The alcohol-containing distillation solution is a solution containing alcohol obtained by a distillation operation, and may be, for example, alcohol for raw materials. Distillation of spirits, whiskey, brandy, wokka, lamb, tequila, gin, shochu, etc. Alcohol and the like can be used.

The lactic acid bacterium growth inhibitor for beer-like effervescent beverages according to the present invention (hereinafter, may be referred to as "lactic acid bacterium growth inhibitor according to the present invention") is characterized by containing polylysine as an active ingredient. Polylysine has high growth inhibitory activity against lactic acid bacteria. Therefore, polylysine is very useful as an antibacterial agent used in beer-like effervescent beverages in which lactic acid bacteria are the main harmful bacteria. Further, polylysine is known to show growth inhibitory activity against both lactic acid bacteria and yeast, but the growth inhibitory activity against lactic acid bacteria is stronger. Therefore, even in a fermented beer-like effervescent beverage, polylysine has a sufficient antibacterial action against lactic acid bacteria, but has no growth inhibitory action against yeast or is contained in a concentration at which the action is weak. This makes it possible to suppress contamination due to the growth of lactic acid bacteria while suppressing poor fermentation and resulting deterioration in quality.

The polylysine used in the present invention may be one extracted from a natural product, one produced by a transforming microorganism, or one chemically synthesized. For example, polylysine can be extracted from this microorganism because it is produced by Streptomyces spp. As the polylysine used in the present invention, commercially available ones can also be used. The commercially available polylysine may be a refined product of polylysine, or may be an additive for foods and drinks containing an active ingredient other than polylysine.

By adding the lactic acid bacterium growth inhibitor according to the present invention to a beer-like effervescent beverage, the growth of lactic acid bacteria in the beer-like effervescent beverage can be suppressed. Lactic acid bacteria are the main harmful bacteria in beer-like sparkling beverages, and if the growth of lactic acid bacteria can be sufficiently suppressed, the risk of microorganisms can be significantly reduced. The amount of the lactic acid bacterium growth inhibitor added to the beer-like effervescent beverage may be an amount sufficient to obtain the effect of suppressing the growth of lactic acid bacteria in the beverage (lactic acid bacterium suppression effect). For example, the content of polylysine in the beverage of the lactic acid bacterium growth inhibitor is preferably 0.0001% (w / v) or more, more preferably 0.0001 to 0.001% (w / v), and even more preferably. By setting it to 0.0002 to 0.0005% (w / v), more preferably 0.0002 to 0.0004% (w / v), it is possible to sufficiently suppress lactic acid bacteria while suppressing quality deterioration due to poor fermentation. The effect can be obtained. The content of polylysine in beer-like effervescent beverages and the like can be measured by a general method used when quantifying a specific protein in a beverage. Examples of the method include high performance liquid chromatography (HPLC) or the like in which the α-amino group of polylysine in a sample is derivatized and then hydrolyzed to produce α-dansyl lysine (α-DNS-Lys). (Hishiyama et al., "Quantification of ε-polylysine in food by pre-column-dancil derivatization high performance liquid chromatography", Journal of Japan Society for Food Science and Technology, October 1996, Vol. 43, No. 10, (Pages 1105-1109).

The lactic acid bacterium growth inhibitor according to the present invention has a high lactic acid bacterium growth inhibitory effect. Therefore, the lactic acid bacterium growth inhibitor according to the present invention particularly comprises a beer-like effervescent beverage not made from hops, that is, an antibacterial component derived from hops. It can be suitably used for beer-like effervescent beverages that do not contain it. By adding the lactic acid bacterium growth inhibitor, the risk of microorganisms can be sufficiently reduced even in a beer-like sparkling beverage that does not use hops as a raw material.

Polylysine also exhibits a high growth inhibitory effect on so-called hop-resistant lactic acid bacteria. Therefore, by using the lactic acid bacterium growth inhibitor according to the present invention, a beer-like effervescent beverage made from hops can also exhibit a sufficient lactic acid bacterium inhibitory effect, and the microbial risk can be sufficiently reduced.

The beer-like effervescent beverage according to the present invention can be produced in the same manner as a general fermented beer-like effervescent beverage or non-fermented beer-like effervescent beverage except that it contains polylysine. Therefore, a method for producing a general fermented beer-like sparkling beverage and a non-fermented beer-like sparkling beverage will be described.

The fermented beer-like effervescent beverage produced through the fermentation step can generally be produced by the steps of preparation (preparation of fermentation raw material liquid), fermentation, storage of sake, and filtration.

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

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 starch such as a grain raw material is saccharified to prepare a sugar solution. As the raw material of the sugar solution, only the grain raw material may be used, only the sugar raw material may be used, or both may be mixed and used. Examples of the grain raw material include barley and wheat, wheat such as malt thereof, beans such as rice, corn and soybean, and potatoes. The grain raw material can be used as a grain syrup, a grain extract, or the like, but is preferably used as a crushed grain product obtained by crushing. The crushing treatment of grains can be carried out by a conventional method. The crushed grain may be a crushed malt, cornstarch, corn grits, or the like, which has been subjected to a treatment usually performed before and after the crushing treatment. In the present invention, the crushed grain product used is preferably a crushed malt product. By using the crushed malt product, it is possible to produce a fermented beer-like sparkling beverage having a clearer beer-like character. The crushed malt product may be any crushed barley, for example, Nijo barley, which is germinated by a conventional method, dried, and then crushed to a predetermined particle size. Further, the grain raw material used in the present invention may be one kind of grain raw material or a mixture of a plurality of kinds of grain raw materials. For example, crushed malt may be used as the main raw material, and crushed rice or corn may be used as the auxiliary raw material. Examples of the sugar raw material include sugars such as liquid sugar.

A grain raw material or an auxiliary raw material other than water may be added to the mixture. Examples of the auxiliary raw material include hops, dietary fiber, yeast extract, fruit juice, bitterness, coloring agents, herbs, and fragrances. Further, if necessary, a saccharifying enzyme such as α-amylase, glucoamylase and pullulanase, and an enzyme preparation such as protease can be added.

The saccharification treatment is carried out using an enzyme derived from a grain raw material or the like or an enzyme added separately. The temperature and time during the saccharification process are determined by the type of grain raw material used, the ratio of the grain raw material to the total fermentation raw material, the type and amount of the added enzyme, the quality of the target fermented beer-like effervescent beverage, etc. It will be adjusted as appropriate in consideration. For example, the saccharification treatment can be carried out by a conventional method, such as holding a mixture containing a grain raw material or the like at 35 to 70 ° C. for 20 to 90 minutes.

By boiling the sugar solution obtained after the saccharification treatment, a broth (boiled product of the sugar solution) can be prepared. It is preferable that the sugar solution is filtered before the boiling treatment and the obtained filtrate is boiled. Further, instead of the filtrate of this sugar solution, a malt extract to which warm water is added may be used and boiled. The boiling method and its conditions can be appropriately determined.

A fermented beer-like sparkling beverage having a desired flavor can be produced by appropriately adding herbs or the like before or during the boiling treatment. For example, by adding hops before or during the boiling treatment and boiling the hops in the presence of the hops, the flavor / aroma components of the hops, particularly the bitterness components, can be efficiently boiled. The amount of hops added, the mode of addition (for example, addition in several portions) and the boiling conditions can be appropriately determined.

The broth obtained after the boiling treatment contains meals such as proteins produced by precipitation. Therefore, at least a part of solids such as lees is removed from the broth. The debris may be removed by any solid-liquid separation treatment, but generally, the precipitate is removed using a tank called a whirlpool. The temperature of the broth at this time may be 15 ° C. or higher, and is generally about 50 to 100 ° C. After removing the dregs, the broth (filtrate) is cooled to an appropriate fermentation temperature with a plate cooler or the like. The broth after removing the lees becomes the fermentation raw material liquid.

Next, as a fermentation step, yeast is inoculated into the cooled fermentation raw material solution to perform 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 adjusting to a desired extract concentration. The yeast used for fermentation is not particularly limited, and usually, yeast used for producing alcoholic beverages can be appropriately selected and used. Top-fermenting yeast may be used, or bottom-fermenting yeast may be used, but bottom-fermenting yeast is preferable because it can be easily applied to large-scale brewing facilities.

Further, as a liquor storage step, the obtained fermented liquor is aged in a liquor storage tank, stored and stabilized under low temperature conditions of about 0 ° C., and then the ripened fermented liquor is filtered as a filtration step. , Yeast and insoluble proteins in the temperature range can be removed to obtain the desired fermented beer-like sparkling beverage. The filtration treatment may be a method capable of removing yeast by filtration, and examples thereof include diatomaceous earth filtration and filter filtration with a filter having an average pore size of about 0.4 to 1.0 μm.

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, may be added to the fermentation liquid in the fermentation step, or may be added to the fermentation liquid in the sake storage step. Since polylysine retains the effect of suppressing lactic acid bacteria even after the heat treatment, it may be added before the boiling treatment in the preparation step or during the boiling treatment.

The pH of the fermentation raw material liquid is adjusted to 5.0 to 6.0 in order to facilitate the precipitation of heat-coagulable proteins and to bring the pH suitable for alcoholic fermentation. In addition, the pH becomes 4.0 to 4.5 after fermentation. Polylysine can suppress the growth of lactic acid bacteria in the pH range of 4.0 to 6.0, preferably 4.0 to 5.5.

A non-fermented beer-like effervescent beverage produced without undergoing a fermentation step can generally be produced by a method (formulation method) of mixing each raw material. For example, specifically, it can be produced by a compounding step of preparing a compounding solution by mixing each raw material and a gas introduction step of adding carbon dioxide gas to the obtained compounding solution.

First, in the compounding step, a compounding solution is prepared by mixing the raw materials. In the compounding step, it is preferable to prepare a compounding solution in which all the raw materials other than carbon dioxide are mixed. The order in which each raw material is mixed is not particularly limited. All the raw materials may be added to the raw material water at the same time, or the raw materials remaining after dissolving the previously added raw materials may be added in sequence. Further, for example, a solid (for example, powder or granular) raw material and alcohol may be mixed with the raw material water, and the solid raw material may be prepared as an aqueous solution in advance, and these aqueous solutions, alcohol, and raw material water may be added if necessary. You may mix.

Examples of the raw material include bitterness, acidulant, sweetener, caramel color, flavor, ethanol (raw alcohol), emulsifier, polysaccharide, water-soluble dietary fiber, protein or its decomposition product.

Examples of the acidulant include lactic acid, citric acid, gluconic acid, tartaric acid, malic acid, succinic acid, phosphoric acid, adipic acid, fumaric acid and the like. Only one kind of these acidulants may be used, or two or more kinds may be used in combination.

Bitterness agents include hops, iso-alpha acids, tetra-iso-alpha acids, β-acid oxides, magnesium salts, calcium salts, naringin, quasin, quinine, momordecine, quercitrin, theobromine, caffeine, bitter gourd, senburi tea, and Kuding. Examples thereof include tea, bitter hop extract, gentiana extract, and quinine extract. Only one kind of these bitterness agents may be used, or two or more kinds thereof may be used in combination.

The hops used as a raw material may be raw hops, dried hops, hop pellets, or processed hops. The processed hop product may be a hop extract obtained by extracting the bitterness component from the hop, and is a processed hop product containing an isometric component of the bitterness component in the hop such as isometric hop extract, tetrahydroisohumulone, and hexahydroisohumulone. May be.

Examples of the sweetener include sucrose, glucose, fructose, high fructose corn syrup, and high-sweetness sweeteners. Examples of high-sweetness sweeteners include aspartame, sucralose, acesulfame potassium, neotame, stevia, and enzyme-treated stevia. Only one kind of these sweeteners may be used, or two or more kinds may be used in combination.

Examples of the flavoring agent include beer extract, beer flavoring, hop flavoring and the like. Only one kind of these flavoring agents may be used, or two or more kinds thereof may be used in combination.

Examples of the emulsifier include polyglycerin fatty acid ester, glycerin fatty acid ester, sucrose fatty acid ester, polypropylene glycol fatty acid ester, sorbitan fatty acid ester, polysorbate and the like.

Examples of the polysaccharide include starch, dextrin and the like. Dextrin is a carbohydrate obtained by hydrolyzing starch and is larger than oligosaccharide (a carbohydrate obtained by polymerizing about 3 to 10 monosaccharides). Only one type of these polysaccharides may be used, or two or more types may be used in combination.

Water-soluble dietary fiber means carbohydrates that are soluble in water and are indigestible or difficult to digest by human digestive enzymes. Examples of the water-soluble dietary fiber include soybean dietary fiber (soluble soybean polysaccharide), polydextrose, indigestible dextrin, galactomannan, inulin, guar gum decomposition product, pectin, and gum arabic. Only one kind of these water-soluble dietary fibers may be used, or two or more kinds may be used in combination.

When an insoluble matter is generated in the prepared liquid prepared in the blending step, it is preferable to perform a treatment for removing the insoluble matter such as filtration on the blended liquid before the gas introduction step. The insoluble matter removing treatment is not particularly limited, and can be carried out by a method usually used in the art such as a filtration method and a centrifugation method. In the present invention, the insoluble matter is preferably removed by filtration, and more preferably removed by diatomaceous earth filtration.

Next, as a gas introduction step, carbon dioxide gas is added to the blending solution obtained in the blending step. As a result, a non-fermented beer-like sparkling beverage is obtained. By adding carbonic acid, the same refreshing feeling as beer is given. The carbon dioxide gas can be added by a conventional method. For example, the blended solution obtained in the blending step and carbonated water may be mixed, and carbon dioxide gas may be directly added to the blended solution obtained in the blending step to dissolve the mixture.

After adding carbon dioxide gas, the obtained non-fermented beer-like effervescent beverage may be further subjected to a treatment for removing insoluble matter such as filtration. The insoluble matter removing treatment is not particularly limited, and can be carried out by a method usually used in the art.

Polylysine can be added as a raw material in any step of the manufacturing process of a non-fermented beer-like sparkling beverage. For example, in the compounding step, it may be added to the compounding solution together with other raw materials, or may be added after introducing carbon dioxide gas and before or after the insoluble matter removing treatment.

When the effervescent beverage according to the present invention is a packaged beverage, the container for filling the effervescent beverage according to the present invention is not particularly limited. Specific examples thereof include glass bottles, cans, and flexible containers. Examples of the flexible container include a container formed by molding a flexible resin such as PE (polyethylene), PP (polypropylene), EVOH (ethylene / vinyl alcohol copolymer), and PET (polyethylene terephthalate). The flexible container may be made of a single layer resin or may be made of a multilayer resin.

Next, the present invention will be described in more detail with reference to Examples and Reference Examples, but the present invention is not limited to the following Examples and the like.

[Reference example 1]
Saccharomyces pastorianus (hereinafter, "S. pastorianus") and an environmentally isolated strain of lactic acid bacteria (Lactobacillus brevis: hereinafter, "L. brevis") were prepared with hop-containing wort and without hops. The proliferative property in wort was examined.

<Preculture of microorganisms and preparation of bacterial solution>
S. pastorianus was inoculated on CP-added potato dextrose agar medium (manufactured by Eiken Chemical Co., Ltd.), and L. brevis was inoculated on MRS liquid medium (manufactured by de Man, Rogosa and Sharpe, manufactured by Merck) at 25 ° C. for 3 Inoculated for days.
The pre-cultured cells, those suspended in sterile saline so that the number of bacteria per 1mL of about 10 ^5, and the bacterial solution.

<Preparation of wort>
A mixture of 40 kg of malt and 160 L of water was held at 58 ° C. for 20 minutes for proteolysis, and then held at 64 ° C. for 20 minutes to saccharify the malt-derived components. The saccharified mixture was filtered to give hop-free clear wort.

The obtained wort was sterilized by heating in an autoclave (121 ° C., 15 minutes), and then the trove was removed with a filter paper. The removed wort was adjusted to pH 5.25 ± 0.05 with 5N HCl or NaOH, and then sterilized by a filter (pore size 0.45 μm) to obtain hop-free wort.
The wort was prepared in the same manner except that hops were added during heat sterilization in an autoclave, and the wort containing hops was sterilized by a filter.

<Culture of microorganisms>
The free hop wort or hop containing wort, S. The pastorianus or L. brevis, seeded such that the 96-well plates to 10 ³ cells per well / 100 [mu] L, and incubated at 25 ° C., over time the absorbance of 600nm Measured to. Absorbance was measured using a "VICTOR Nivo Multimode Plate Reader" (manufacturer: PerkinElmer, S / N: HH35L3517059, excitation filter: 600/10 nm). A microbial uninoculated sample was used as a background measurement.

The measurement results of the wort to which L. brevis was added are shown in FIG. 1 (A), and the measurement results of the wort to which S. pastorianus was added are shown in FIG. 1 (B). As a result, S. pastorianus was observed to grow in all worts, while L. brevis was confirmed to grow in hop-free wort with increased absorbance, but not in hop-containing worts. ..

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

<Preparation of polylysine raw material>
The polylysine is an aqueous solution (10% (w / v) polylysine solution) adjusted with pure water so as to have a final concentration of 10% (w / v), and the final concentration of polylysine is 0.0010% (w / v). As described above, the mixture was diluted with hop-free wort, adjusted to pH 5.25 ± 0.05 with hydrochloric acid or lactic acid, and then sterilized by a filter (pore size 0.45 μm). By further diluting the obtained 0.0010% (w / v) polylysine solution with hop-free wheat juice, the final concentrations of polylysine were 0.00001% (w / v) and 0.00002% (w / v). ), 0.00004% (w / v), 0.00006% (w / v), 0.00008% (w / v) polylysine solutions were prepared.
The sample used for the evaluation of heat resistance was prepared in the same manner using a 10% (w / v) polylysine solution heated at 100 ° C. for 30 minutes.

<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, 90 μL of the polylysine raw material solution adjusted to each concentration was added, and the temperature was 25 ° C. for 7 days. The cells were cultured under anaerobic conditions. The viable cell count (test cell count) of the inoculated bacterial solution was measured by a plate smear culture method using CP-added potato dextrose agar medium or MRS agar medium, and was converted into the viable cell count of the inoculated bacterial solution.
In addition, the solution in each well was measured for absorbance at 600 nm over time in the same manner as in Reference Example 1, and the growth of microorganisms was examined.

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 FIG. 2 (A) from the heat-treated 10% (w / v) polylysine solution. The results of the prepared polylysine solution are shown in FIG. 2 (B), respectively. Further, among the samples in which S. pastorianus was sown, the results of the polylysine solution prepared from the unheated 10% (w / v) polylysine solution are shown in FIG. 3 (A), and the heat-treated 10% (w / v) polylysine was obtained. The results of the polylysine solution prepared from the solution are shown in FIG. 3 (B), respectively.

As a result, it was confirmed that in all the samples in which L. brevis was sown, the increase in absorbance was suppressed and the growth of L. brevis was suppressed as compared with the result of the hop-free wort of Reference Example 1. It was. In addition, an increase in absorbance was confirmed in all the samples in which S. pastorianus was sown, confirming that S. pastorianus could proliferate.

Among them, in the solution to which the polylysine concentration was added 0.0004% (w / v), the growth of L. brevis was hardly confirmed in either the unheated polylysine or the heat-treated polylysine ( 2 (A) and 2 (B)), S. pastorianus showed sufficient proliferation (FIGS. 3 (A) and (B)). From these results, it was confirmed that the growth of lactic acid bacteria can be suppressed while suppressing the influence on fermentation by containing an appropriate amount of polylysine.

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

<Adjustment of polylysine raw material>
For the adjustment of polylysine raw materials, commercially available polylysine preparations for food additives (product name: "Sunkeeper No.381", manufactured by Sanei Gensha, composition: polylysine 50% (w / w), glycerin 50% (w / w)) )It was used. The polylysine preparation was added to hop-free wort so that the final concentration of polylysine was 0.0005, 0.00125, 0.0025, or 0.005% (w / v), and this was used as a raw material for polylysine. ..

<Inoculation of test bacteria>
Example 1 was suspended in non-hop wort as the number of bacteria is approximately 40 × 10 ⁸ cells per 1mL Cells of pre-cultured S. pastorianus were similarly used as a cell solution. 10 mL of the bacterial solution was inoculated into an Erlenmeyer flask, 190 mL of polylysine raw material adjusted to each concentration was added, mixed, fermented, and cultured at 10.5 ° C. for 8 days under aerobic conditions.

In alcoholic fermentation, 2 molecules of ethanol and 2 molecules of carbon dioxide are produced from 1 molecule of sugar [C ₆ H ₁₂ O ₆ (180 g) → 2 C ₂ H ₅ OH (46 g × 2) + 2CO ₂ (44 g × 2) )]. Therefore, the concentration (g / L) of alcohol produced by alcoholic fermentation can be expressed by the following formula (1). Further, since the specific gravity of ethanol is 0.789 g / mL, it is converted into the volume concentration by the following formula (2). Therefore, the amount of carbon dioxide reduction was determined by measuring the weight of the Erlenmeyer flask during culture, and the alcohol concentration (% (v / v)) produced was estimated.

Formula (1): [Alcohol concentration of fermented liquid (g / L)] = ((46/44) x [carbon dioxide reduction amount (g)]) / [culture liquid amount (L)]
Formula (2): [Alcohol concentration of fermented liquid (% (v / v))] = (0.1325 x [carbon dioxide gas reduction amount (g)]) / [culture liquid amount (L)]

Table 1 shows the measurement results of the alcohol concentration (% (v / v)) of each reaction solution. The higher the final concentration of polylysine in the fermentation broth, the lower the amount of alcohol produced. However, even if the final concentration of polylysine was 0.005%, the alcohol concentration of the fermentation broth was about 0.7% (v / v), and it was confirmed that alcohol production was carried out without any problem.

[Example 3]
Alcoholic fermentation was carried out by mass culture in the presence of polylysine, and the obtained fermented beer-like effervescent beverage was sensory evaluated. As S. pastorianus and L. brevis, those used in Reference Example 1 were used.

<Manufacturing of fermented beer-like sparkling beverage>
22 kg of cornstarch and 66 kg of water were mixed and heated to swell (swelling starch). Next, a mixture of 44 kg of malt and 110 kg of water was held at 55 ° C. for 30 minutes for proteolysis. The swollen starch was then added to the mixture and held at 64.5 ° C. for 50 minutes to saccharify the malt and cornstarch-derived components and filter to give a clear wort. Then, the wort was boiled for 80 minutes without adding hops, and the prepared hot water was added. To the obtained wort after boiling, the polylysine preparation used in Example 2 was added and stirred so that the final concentration of polylysine was 0.0005% (w / v), and after removing the thermal coagulant, about 8 Cooled to ° C. The cooled hop-free wort was transferred to a fermentation tank and added so that the numbers of S. pastorianus and L. brevis were 20 × 10 ⁶ and 1.0 × 10 ⁵ per mL, respectively. It was cultured at 5 ° C.

Bacterial concentration and extract consumption of S. pastorianus during fermentation were measured. The measurement results are shown in FIG. The results of the control sample obtained by alcoholic fermentation of wort without polylysine are shown in FIG. 4 (A), and the results of the test sample obtained by alcoholic fermentation of wort having a final concentration of polylysine of 0.0005% (w / v) are shown in FIG. 4 (B) shows each. In addition, sampling was performed every 7 days of fermentation, and the viable cell count of L. brevis was observed. The measurement results are shown in Table 2.

As shown in FIG. 4, in wort containing a final polylysine concentration of 0.0005% (w / v), S. pastorianus proliferated as in the control and consumed the extract. In addition, as shown in Table 2, the viable amount of L. brevis gradually decreased during the fermentation and aging period of wort containing a final polylysine concentration of 0.0005% (w / v).

<Sensory evaluation>
The obtained test sample and control sample were first clarified by filtration, diluted 1.2 times, and then filled in a container. Next, in order to investigate whether or not there was a change in flavor due to the addition of polylysine, both samples were subjected to sensory evaluation by a three-point test method. The sensory evaluation was carried out 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.

As shown in Table 3, of the 11 panels, 2 were the ones who correctly identified the test sample and the control sample, and no significant difference was observed between the two samples. That is, even if alcoholic fermentation was carried out in the presence of polylysine, a fermented beer-like effervescent beverage having an excellent flavor as in the absence of polylysine could be produced.

[Example 4]
The proliferative potential of two yeasts other than S. pastorianus in the presence of polylysine was investigated. As yeast, Saccharomyces cerevisiae (hereinafter, "S. cerevisiae") and Saccharomyces bayanus (hereinafter, "S. bayanus") were used.

Using S. cerevisiae or S. bayanus instead of S. pastorianus as yeast, the polylysine raw material solution prepared in Example 1 or the commercially available polylysine preparation for food additives used in Example 2 (product name: "Product name:" In the same manner as in Example 1 except that "Sunkeeper No. 381" (manufactured by Sanei Gensha) was used, the yeast solution was mixed with polylysine at a final concentration of 0.0005% (w / v) in a 96-well plate well. And cultured at 25 ° C. for 7 days under anaerobic conditions. The absorbance at 600 nm of each well was measured over time to examine the growth of microorganisms.

The measurement result of the absorbance at 600 nm of each well is shown in FIG. The change in absorbance between S. cerevisiae (“S. cerevisiae / polylysine preparation” in the figure) to which the polylysine preparation was added and S. cerevisiae (“S. cerevisiae / polylysine raw material liquid” in the figure) to which the polylysine raw material solution was added was almost the same. Similarly, S. cerevisiae showed sufficient proliferation in the presence of 0.0005% (w / v) polylysine. Similarly, the absorbance of S. bayanus (“S. bayanus / polylysine preparation” in the figure) to which the polylysine preparation was added and S. bayanus (“S. bayanus / polylysine raw material liquid” in the figure) to which the polylysine raw material solution was added. The changes were similar and S. bayanus also showed sufficient proliferation in the presence of 0.0005% (w / v) polylysine.

Claims (8)

A lactic acid bacterium growth inhibitor for beer-like effervescent beverages, which comprises polylysine as an active ingredient. A beer-like effervescent beverage characterized by containing polylysine. The beer-like effervescent beverage according to claim 2, wherein the content of polylysine is 0.0001% (w / v) or more. The beer-like sparkling beverage according to claim 2 or 3, which is not made from hops. The beer-like effervescent beverage according to any one of claims 2 to 4, which is a fermented beer-like effervescent beverage. A method for producing a beer-like effervescent beverage, which comprises using polylysine as a raw material. The method for producing a beer-like effervescent beverage according to claim 6, wherein hops are not used as a raw material. The method for producing a beer-like effervescent beverage according to claim 6 or 7, wherein the produced beer-like effervescent beverage has a polylysine content of 0.0001% (w / v) or more.

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