JP3288993B2 - Antifungal agent effective against drug-resistant fungi - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

[0001] The present invention relates to a drug-resistant fungus comprising an organic solvent extract of a licorice or licorice water extract residue,
In particular, the genus Arthrinium and Ketomium (Cha)
etomium) which is effective against fungi of the genus Fungi.

[0002]

2. Description of the Related Art Licorice is conventionally known as a crude drug, and is currently mainly used as a raw material for food sweeteners, pharmaceuticals and quasi-drugs. In particular, glycyrrhizin and glycyrrhetinic acid, which are water-soluble components, have been widely used in foods, pharmaceuticals, cosmetics, etc. because of their excellent pharmacological actions such as anti-inflammatory action, anti-ulcer action, and anti-allergic action. Furthermore, licorice contains many flavonoids in addition to glycyrrhizin, and it is known that the fraction obtained by extracting licorice with an organic solvent contains an antibacterial substance against bacteria.

[0003] Organic solvent extracts of licorice include gram-positive bacteria [Staphylococcus aureus (JP-A-59-46210),
Bacillus subtilis (JP-A-60-233015), Stre
ptococcus mutans (JP-A-63-145208), Bacill
us cereus, Bacillus licheniformis, Lactobacillus c
asei, Lactobacillus arabinosus, Streptococcus faec
alis (Monthly Food Chemical, p. 94 (1989, April)) is known to have an excellent growth inhibitory effect.
However, the antibacterial effect is very weak against Gram-negative bacteria (JP-A-63-24489, JP-A-63-145208, and JP-A-8-295632).

On the other hand, there is very little knowledge about the antibacterial activity of an organic solvent extract of licorice against mold, and the fungicide whose antibacterial effect has been clearly reported is Penicillium chrysogenum.
(JP-A-59-46210) and Mucor pusillus (JP-A-59-46210).
63-145208). On the other hand,
Aspergillus niger, Penicillium citrinum, Rhizopus
It is also reported that nigricans and Fusarium graminearum have almost no antibacterial effect (Food and Packaging, Vol. 24, pp. 53-65, 1992; JP-A-63-145208). As the above findings indicate, at present, the antibacterial spectrum of the organic solvent extract of licorice against fungi cannot be predicted at all.

[0005] By the way, a wide variety of soft drinks are manufactured in the present beverage industry, and the market scale thereof is steadily expanding. Low acid beverages (soft drinks other than mineral water, frozen fruit beverages, and fruit juices for raw materials that have a pH of 4.6 or more and have a water activity of more than 0.94) are playing a leading role, Among them, tea-based beverages have shown remarkable growth, helped by the growing functionality and health of consumers. Concerning soft drink containers, with the abolition of regulations on small plastic bottles, which were voluntarily regulated in the soft drink industry (April 1996), consumer needs have been able to recap from canned beverages. The selection of small plastic bottles has diversified, and the market for soft drinks in small plastic bottles is expanding rapidly.

Green tea, oolong tea, black tea, milk tea,
Low-acid beverages such as coffee, milk coffee, barley tea, mixed tea (blend tea), cocoa, and milkshake tend to grow more easily than carbonated beverages and acidic beverages. is necessary. In fact, according to the production standards of soft drinks described in the Food Sanitation Law, sterilizing conditions for low acid beverages (120
Acidic beverages (those having a pH of less than 4.0: 65 ° C, 10 minutes or equivalent or more, and those having a pH of 4.0 to less than 4.6): 85
(30 ° C., 30 minutes or equivalent or more).

Therefore, especially in the case of a low-acid beverage,
Except for the most popular canned soft drinks at present, the use of non-heat-resistant containers such as plastic bottles, bag-in-boxes or paper containers requires that the containers cannot withstand the sterilization conditions required by the aforementioned Food Sanitation Law. A process is used in which soft drinks are sterilized by heating under necessary conditions in advance, cooled, and then filled into a non-heat-resistant container.

[0008] In such a manufacturing process, the beverage after sterilization must be filled and packaged in a sterile container in a sterile environment in order to prevent contamination by microorganisms. Such an entire manufacturing system is called an aseptic filling and packaging system. When the aseptic filling and packaging system is used, containers and caps that are not heat-resistant are used in terms of manufacturing costs. For this reason, chemicals such as sodium hypochlorite, hydrogen peroxide, peracetic acid, and ozone water are used for sterilization of containers and caps.

[0009]

However, recently, microorganisms having resistance to these sterilizing agents have been found, and there is a concern that aseptic-filled low-acid beverages may be degraded and contaminated by the drug-resistant microorganisms. Above all, fungi of the genus Earthrinium and the genus Ketomium are highly resistant to agents used for sterilization of non-heat-resistant containers. Therefore,
Inhibiting the growth of these fungi has become a very important issue in the production of the above beverages. In order to sterilize these fungi, it is necessary to take countermeasures such as increasing the sterilization time, increasing the sterilization temperature and drug concentration, or using multiple sterilizing agents (Beverage Japan, No.
193, 76-78 (January 1998)),
Reduction of production efficiency and cost increase are inevitable, and are not a sufficient solution. Therefore, it is desired to minimize the decrease in production efficiency and the increase in cost and to suppress the growth of drug-resistant fungi, in particular, the fungi of the genus Earthrinium and the genus Ketomium.

[0010]

Means for Solving the Problems The present inventors have proposed an effective and highly safe antibacterial substance which can be directly added to beverages as a method for preventing the deterioration of these drug-resistant fungi in beverages. Set out to search. When imparting antibacterial properties to low-acid beverages, especially non-heat-resistant packaged tea beverages, they must meet the minimum conditions that do not adversely affect the properties of the beverage, such as taste, aroma, light blue, and clarity. Preferably, it is a natural product that can be used as a beverage material if possible.
Further, in addition to not changing the properties of the beverage as described above, in order not to significantly increase the cost of the target beverage, a highly effective antibacterial substance that can exhibit a sufficient antibacterial effect at a low concentration Must.

Therefore, the present inventors have studied the taste, aroma,
It does not adversely affect properties such as light blue and clarity, does not significantly increase the cost of the target beverage, can exhibit sufficient antibacterial effects even at low concentrations, and is derived from natural products that can use antibacterial substances as beverage ingredients As a result of intensive research on searching for highly effective antibacterial substances against fungi-resistant earthlinium and ketomium fungi, the organic solvent extract of licorice or licorice water extraction residue was found to be The present inventors have found that they exhibit a growth inhibitory effect on the above-mentioned drug-resistant bacteria at an extremely low concentration, and have completed the present invention.

That is, according to the present invention, sodium hypochlorite, a sterilizing agent, comprising an organic solvent extract of licorice or a licorice water extraction residue,
It is an effective antifungal agent for fungi resistant to hydrogen, peracetic acid, or ozone water . According to a second aspect of the present invention, there is provided the antifungal agent according to the first aspect, wherein the sterilizing drug-resistant fungus is a fungus belonging to the genus earthrinium and / or the genus ketomium. The present invention according to claim 3 is the antifungal agent according to claim 1 or 2, wherein the organic solvent extract of the licorice or licorice water extraction residue is obtained by extracting the licorice or licorice water extraction residue with ethanol or hydrous ethanol. is there.

[0013]

BEST MODE FOR CARRYING OUT THE INVENTION The licorice used in the present invention belongs to a plant belonging to the genus Glycyrrihiza in the legume family, and has a root,
Any part of the rhizome, leaf, or stem may be used, regardless of whether it is in a raw or dry state, but it is easy to use dry roots and dry rhizomes that are industrially produced glycyrrhizin extraction raw materials. Is also preferred. Licorice is often named after the place of production, and examples include eastern licorice, northeast licorice, Xinjiang licorice, Mongolian licorice, Russian licorice, and Afghan licorice.

The licorice water extraction residue of the present invention is a solid residue obtained by extracting the above licorice with cold, warm, hot or neutral or slightly alkaline cold, warm or hot water, or a combination of these residues. This is a solid residue after extraction, and the residue after extraction may be either water-containing or dry. In order to obtain the fraction having the antifungal activity according to the present invention from the licorice or licorice water extraction residue, various organic solvents may be extracted alone or in combination. Examples of the organic solvent include benzene, toluene, xylene, ethyl ether, methyl ethyl ketone, methyl isobutyl ketone,
Examples include dichloromethane, dichloroethane, chloroform, ethyl acetate, propyl acetate, butyl acetate, acetone, methanol, ethanol, propanol, hydrated methanol, hydrated ethanol, and hydrated propanol. Further, carbon dioxide can be used as a supercritical fluid. Among these organic solvents, it is preferable to use ethanol or hydrous ethanol since there is no problem in the Food Sanitation Law.

The conditions for extracting the substance having an antifungal activity from the licorice or licorice water extraction residue with the above-mentioned organic solvent are not particularly limited. There is a method of adding an organic solvent in an amount of 10 to 10 times and extracting at room temperature while stirring, or a method of extracting by heating to reflux. In addition, each of these methods alone,
Or it is more preferable to perform the operation repeatedly in combination because the extraction efficiency is improved.

After removing the insoluble matter from the obtained extract by centrifugation or filtration, it can be used as it is as a liquid extract or further concentrated by a conventional method. If the extract is dried by an appropriate method, a tan extract powder can be obtained. These organic solvent extracts can be directly used as the antifungal agent of the present invention, but may be appropriately subjected to purification such as deodorization and decolorization as long as the antibacterial effect is not reduced. In this purification step, it is common to use activated carbon, a synthetic adsorption resin, an ion exchange resin and the like.

The organic solvent extract of the licorice or licorice aqueous extract residue, which is the antifungal agent of the present invention, produced by the above method is oil-soluble and does not dissolve even if the powdery extract is added to an aqueous solution as it is. For this reason, it is preferable to dissolve it in a solvent such as acetone, methanol, ethanol, propylene glycol or the like in advance and then add it to the aqueous solution. In particular, in the case of adding to a beverage which is the object of the present invention, the antifungal agent of the present invention is dissolved in a dissolving agent permitted to be used in the Food Sanitation Law, for example, ethanol, hydrous ethanol, propylene glycol or a mixture thereof. Is appropriate.

[0018] In addition, low-acid beverages include barley tea, mixed tea, green tea, oolong tea and black tea-based beverages, except for non-transparent beverages such as coffee with milk, black tea with milk, cocoa and milkshake. Since the beverage is a certain beverage, when the antifungal agent of the present invention is dissolved in the above-mentioned dissolving agent and then added to the beverage, it may be dispersed but impair the transparency. In order to prevent this, after dissolving the organic solvent extract of the licorice or licorice water extraction residue in the above-mentioned dissolving agent, a surfactant that is recognized as being added to food, for example, glycerin fatty acid ester, propylene glycol fatty acid ester, Sorbitan fatty acid ester, sucrose fatty acid ester, soy lecithin, vegetable saponin, etc. can be added alone or in combination to form a water-soluble antifungal agent, so that the transparency can be maintained even when added to beverages. .
It is also possible to add the above-mentioned surfactant to a beverage in advance and to dissolve it by adding an organic solvent extract of licorice dissolved in a dissolving agent.

When the antifungal agent of the present invention is added to a low-acid beverage, it is added to a container such as an extraction, filtration, concentration, blending, sterilization, or aseptic surge tank during the production process of the low-acid beverage. If the previous process,
The antifungal agent of the present invention may be added at any stage, but in order to uniformly disperse or dissolve the antifungal agent of the present invention, it is preferable to add the antifungal agent at a stage before the sterilization step, and to adjust the taste, aroma, concentration, etc. of the beverage. It is particularly preferred to add in stages.

In the present invention, the low-acid beverage refers to a beverage having a pH of 4.6 or more and a water activity exceeding 0.94 among soft drinks other than mineral water, frozen fruit drinks and fruit juice for raw materials. For example, barley tea, mixed tea,
Green tea, black tea, oolong tea, coffee, cocoa, etc. Here, mixed tea (blend tea) refers to leaves of various plants,
Water extraction using two or more types of stalks, roots, nuts, flowers, bark, etc. as raw materials, generally means beverages extracted with hot water or hot water, and various plant extracts, functional food materials, vitamins, etc. Includes formulated beverages.

Here, examples of the raw materials used include barley, wheat, pigeon barley, green tea, black tea, oolong tea, puar tea, wrapping tea, houjicha, brown rice tea, matcha tea, dokudami, hub grass, soybean, red bean. , Kombu, pong, reishi, wolfberry leaf, wolfberry, juniper leaf, rooibos tea, persimmon leaf, kumasasa, amachazuru, moloheiya, kidachi aloe, jasmine, gymnema, plantain, tangerine peel Guava tea, mulberry leaves,
Buckwheat berries, loquat leaves, safflower, cinnamon, pepper, millet, millet, licorice, chrysanthemum, ginseng, ukogi, shiitake, shiso leaves, sweet tea,
Rosemary, hibiscus, lemon balm, basil,
Sage, lemongrass, mint, chamomile, turmeric,
Tarago, oregano, thyme, coriander, etc.
In addition, tea, coffee and cocoa, whether sugar-free or sweetened, include milk-containing black tea, milk-containing coffee, milk-containing cocoa, and the like.

The low-acid beverage of the present invention comprising an antifungal agent effective against the fungi of the genus Arthrinium and the genus Ketomium is a hypochlorous acid in which the above two fungi are used for sterilization of a non-heat-resistant container. Because it is a microorganism that is resistant to sterilizing agents such as soda, hydrogen peroxide, peracetic acid, ozone water,
It is suitable for beverages to be filled by an aseptic filling and packaging system into a non-heat-resistant container which has been sterilized with a drug in advance. Earthlinium and ketomium fungi are
Since it is not a mold having heat resistance, there is almost no need to add the antifungal agent of the present invention to a canned beverage which is sterilized at a high temperature by retort or the like after filling. The term “non-heat-resistant container” as used herein means a container that loses its original performance, particularly its sealing property and shape under heating conditions of 95 ° C. for 10 minutes or more. .

The aseptic filling in the present invention will be described as follows. That is, when using a non-heat-resistant container that cannot withstand the sterilization conditions required by the Food Sanitation Law, heat sterilize the soft drink in advance under the required conditions, cool it, and then cool it. Is used. In the case of passing through such a manufacturing process, the beverage after the sterilization treatment is filled and packaged in a sterilized container with a chemical agent under an aseptic environment in order to prevent contamination of microorganisms. Such a manufacturing method for manufacturing a beverage in a sterile environment after sterilizing the beverage is referred to as an aseptic filling and packaging system.

When an aseptic filling and packaging system is used, containers and caps that are not heat-resistant are used. For this reason, in the sterilization treatment of containers and caps, drugs used for sterilization and sterilization of microorganisms are generally used in a food production process, and specific examples include:
Examples thereof include sodium hypochlorite, hydrogen peroxide, peracetic acid, and ozone water. Generally, peracetic acid (including a peracetic acid preparation) or ozone water is used. Also, the sterile environment here, work environment and clean room, American Space Administration (NASA) particles class 10,000 (the grain diameter of 0.5 microns or more air standards per cubic foot 10, 000
, 65 particles of 5 microns or more and 1 fallen bacterium
An environment (represented as class 10,000, class 100, etc.) that is less than or equal to 6,000 per week per square foot and does not indicate complete sterility.

Sterilization at the time of production of a low-acid beverage is performed at 120 ° C. for 4 minutes or at least equivalent thereto (the value F used for evaluating the heat sterilization effect of food) specified in the production standard of the Food Sanitation Law. ₀ and 3.1 or more), but in an aseptic filling and packaging system, high-temperature, short-time sterilization is suitable in order to minimize production efficiency and beverage flavor as much as possible. Generally, 120
Or F ₀ value of 4 to about 40 at 0.99 ° C., may be sterilized under the conditions F ₀ value becomes 5 to 30 about a normal 130 to 140 ° C.. In the process after sterilization, it is necessary to consider the heat resistance of the container. However, the temperature of the sterilized beverage may be cooled to 10 to 70 ° C, usually about 20 to 40 ° C.
The F ₀ value as used herein refers to a heating time (minutes) required to kill microorganisms having a constant concentration at a constant temperature, and is usually 250 F ° (sterilization temperature 121.11 ° C.)
Is defined as the heat lethal time in minutes. The details are described in, for example, "Food Sterilization Engineering", Isao Shibasaki, published by Korin Shoin, pp. 54-153 (1967).

The concentration of the antifungal agent of the present invention added to the low-acid beverage produced by the aseptic filling and packaging system may be an amount capable of suppressing the growth of fungi of the genus Earthrinium and the genus Ketomium. Specifically, the organic solvent extract concentration of licorice is 4 ppm or more, preferably 7.5 p.
pm or more, particularly preferably 12 ppm or more is suitable for exhibiting the growth suppressing effect. The upper limit of the additive concentration is determined in a concentration range that does not change properties such as the flavor (taste, aroma, light blue) of the beverage, and is usually 200.
ppm. Therefore, a general concentration range of 4 to 200 ppm is appropriate, but the added concentration slightly varies depending on the characteristics of the beverage. For example, for coffee or tea containing milk, which has a relatively strong taste and does not need to emphasize clarity, it is 4 to 200 ppm, preferably 7.5 to 1 ppm.
The content is preferably 00 ppm, more preferably 12 to 60 ppm. Further, in barley tea, mixed tea, black tea, green tea, oolong tea and the like, the content is 4 to 100 ppm, preferably 7.5 to 60 ppm, particularly preferably 12 to 30 ppm. The reason for limiting the additive concentration in this way is that if the concentration is lower than 4 ppm, the effect of the antifungal agent of the present invention is remarkably reduced, while if it exceeds 200 ppm, the flavor and properties of the beverage are greatly changed. .

Examples of the non-heat-resistant container used for the low-acid beverage containing the antifungal agent of the present invention include a plastic bottle, a bag-in-box and a paper container. As for plastic bottles, polyethylene terephthalate (PET) bottles are currently most widely used,
There are containers using high-density polyethylene, polypropylene, polystyrene, and the like, and these containers can also be used. Bag in box or
Bag in carton) is a liquid container in which a flexible liquid container is used as the interior and corrugated cardboard or paperboard is used as the exterior, and the both are combined. There are many types of liquid containers, such as a double structure of a plastic film, a liquid container having a multi-layer structure such as nylon / polyethylene / aluminum / polyethylene with a liquid-contacting portion (inner layer) of polyethylene and an outer layer portion, which are used depending on the type of beverage. be able to. In addition, the container has a capacity of about 2 to 20 liters, and can be used for business use. There are many types of paper containers, such as “Revised New Edition / Soft Drinks”.
496 pages (Korin, published in 1989), New Food Industry, 35 (No.4),
71-80 (1993), and any container described therein can be used.

[0028]

EXAMPLES The present invention will be described below with reference to examples and the like , but the present invention is not limited by the description. Example 1 One liter of an 80% aqueous methanol solution was added to 100 g of ground licorice root and stirred at room temperature for 24 hours. The extraction residue was removed by filtration, and the filtrate was concentrated under reduced pressure until the filtrate became 100 to 150 mL. The same amount of chloroform was added to the concentrated solution, and after sufficient stirring, the chloroform layer was separated and collected. This operation was repeated twice more. The obtained chloroform layer was concentrated to dryness under reduced pressure, and further dried under reduced pressure. The dried product was pulverized to obtain licorice organic solvent extract (1) (1.9 g).

Example 2 100 g of ground licorice root was extracted with 1 liter of hot water at 90 ° C. for 1 hour. Solid-liquid separation was performed by filtration, and the filtrate was concentrated under reduced pressure and freeze-dried to obtain 19 g of a licorice hot water extract.
The extraction residue was air-dried at 60 ° C., added with 1 liter of ethanol, and stirred and extracted at 40 ° C. for 24 hours. After solid-liquid separation, the ethanol extract was concentrated under reduced pressure to dryness. This dried product was pulverized and then dried under reduced pressure to remove the remaining ethanol, to obtain 2.6 g of licorice organic solvent extract (2).

Example 3 3 g of commercially available oil-soluble licorice extract P-TD (manufactured by Maruzen Pharmaceutical Co., Ltd.) was mixed with ethanol-propylene glycol (1: 1) 6
After dissolving in 0 mL, 3 g of an emulsifier (trade name: L1695, manufactured by Mitsubishi Chemical Food Corp., sucrose laurate) was added. Distilled water was added to this solution to make the total amount 100 mL, thereby preparing a licorice organic solvent extract preparation.

Test Example 1 Licorice organic solvent extract (1) prepared in Example 1, licorice hot water extract and licorice organic solvent extract (2) prepared in Example 2, commercially available oil-soluble licorice extract P- 0.2 g of each sample of TD (manufactured by Maruzen Pharmaceutical) was placed in a sterilized 15 mL tube, and 10 mL of a 10% aqueous solution of dimethyl sulfoxide (DMSO) was added thereto to dissolve. 5 mL of each sample solution was mixed with a 10% DMSO solution to dilute the sample concentration to 1/2. Repeat this operation 9 times,
A dilution series of the sample was prepared.

Next, a sterilized 1.5% potato dextrose agar medium (manufactured by Merck) was cooled to 60 ° C., and chloramphenicol was added to a final concentration of 10 ppm. After 1 mL of each of the sample solutions having the above concentrations was placed in a sterilized petri dish, 20 mL of a potato dextrose agar medium was added and mixed well. At this time, since the final concentration of the sample solution in the medium is a concentration of 10 steps including the undiluted solution, the concentrations of the licorice extract are 1000, 500, 250, 125, 62.5, 31.3, and 1.3, respectively.
It becomes 15.6, 7.8, 3.9, and 2.0 ppm. In the center of the plate agar medium thus prepared, a spore solution (about 1 × 10 ⁵ ) of Arthrinium saccchari or Chaetomium funicola was added.
(Unit / mL) was added dropwise and left at room temperature for 2 hours.
Then, this was cultured at 25 ° C. for 48 hours, and the minimum growth inhibitory concentration (M
IC). After 72 hours of culture, the growth of these two molds at concentrations below the minimum growth inhibitory concentration was determined by measuring the diameter of the colonies with calipers.
The MIC results for each extract are shown in Table 1 and the MI
Table 2 shows the growth rates of earthlinium and ketomium below C.

[0033]

[Table 1] Table 1

As is clear from the table, the licorice hot water extract did not show a growth-inhibiting effect even at 1000 ppm against the drug-resistant bacteria earthlinium and ketomium. In contrast, the effects of the licorice organic solvent extracts (1) and (2) and the commercially available oil-soluble licorice extract at a very low concentration of 31.3 to 62.5 ppm suppress the growth of the above two types of drug-resistant fungi. Was observed.

[0035]

[Table 2] Table 2

As is evident from Table 2, the growth of the above two drug-resistant fungi at concentrations up to 7.8 ppm was observed in any of the licorice organic solvent extracts (1) and (2) and the oil-soluble licorice extract. Suppressed. In addition, the licorice organic solvent extract (2) and the oil-soluble licorice extract showed a growth inhibitory effect even at an extremely low concentration of 3.9 ppm. Thus, the licorice organic solvent extract (including the oil-soluble licorice extract) was an excellent antifungal agent against the above drug-resistant fungi.

Test Example 2 10 mL of sterile water was added to 10 mL of the licorice organic solvent extract preparation prepared in Example 3 to dilute the sample, and the sample concentration was reduced to 1/2.
And This operation was repeated seven times to prepare a dilution series of the sample. Next, a sterilized 1.5% potato dextrose agar medium (manufactured by Merck) was cooled to 60 ° C., and chloramphenicol was added to a final concentration of 10 ppm. After 1 mL of the sample solution of each concentration described above was placed in a sterilized petri dish, 20 mL of a potato dextrose agar medium was added and mixed well. At this time, since the final concentration of the sample solution in the medium has eight levels including the undiluted solution, the licorice organic solvent extract concentration is 750, 375, 188, 94, 47, 24, 1 respectively.
It becomes 2.6 ppm. In the central part of the plate agar medium thus prepared, a chaetomium fu
nicola) or Arthriniu Sacchari (Arthriniu
5 μL of spore solution (approximately 1 × 10 ⁵ cells / mL)
Was added dropwise and left at room temperature for 2 hours. Then, this is 25
After culturing at 48 ° C for 48 hours, colonies were visually observed to determine the minimum growth inhibitory concentration (MIC). After 96 hours of culture,
The growth of these two molds at concentrations below the minimum growth inhibitory concentration was determined by measuring the diameter of the colonies with calipers. The results are shown in Tables 3 and 4.

[0038]

[Table 3] Table 3

[0039]

[Table 4] Table 4

As is evident from Table 3, the licorice organic solvent extract preparation exhibited an inhibitory effect on growth of the drug-resistant fungi earthlinium and ketomium even at very low concentrations. In addition, this extract preparation exhibited an inhibitory effect on the growth of these two types of drug-resistant fungi even at an extremely low concentration of 6 ppm, and was found to be an excellent antifungal agent (Table 4).

Test Example 3 In a clean bench, commercially available 500 mL PET
The licorice organic solvent extract preparation prepared in Example 3 was mixed with the bottled mixed tea, and the licorice organic solvent extract concentration was 50, respectively.
25, 12.5, 6.25, 0 ppm,
Six mixed teas of each concentration were prepared. Each of these three mixed teas has Ketomium funicola
Or Arthrinium sacccha
5 μL of the spore solution (approximately 1 × 10 ⁵ cells / mL) of ri) was added and sealed. This mixed tea in PET bottle is 25 ℃
After 3 weeks, the growth of these two molds (floating matter) was visually observed. The results are shown in Table 5.

As a result, the above two kinds of molds were 6.25p
Growth was observed up to the added concentration of pm, but 12.5 pp
At a licorice organic solvent extract concentration of m or more, the growth was completely suppressed. Therefore, the licorice organic solvent extract exhibited an effect of suppressing the growth of the above two types of mold at an extremely low concentration even when added to a low-acid beverage in which microorganisms actually proliferate.

[0043]

[Table 5] Table 5 +: Growth of bacteria is recognized-: growth of bacteria is not recognized

Test Example 4 The licorice organic solvent extract preparation prepared in Example 3 was applied to commercially available barley tea and milk tea, respectively, at a licorice extract concentration of 400, 200, 100, 60, 30, 10, 5 ppm.
And the taste of these beverages was sensory evaluated by 10 panelists. The results are shown in Table 6. The evaluation criteria for the flavor are as follows.
It was expressed as the average score of 0 panelists.

Rating 0: Flavor does not change 1: Flavor changes slightly but has no adverse effect 2: Flavor changes but does not negatively affect 3: Flavor has a slight adverse effect 4: Flavor has a negative effect

[0046]

[Table 6] Table 6

As shown in Table 6, the upper limit concentration which does not adversely affect the flavor is 200 ppm for milk tea,
In barley tea, it was 100 ppm. However, in the case of barley tea, considering clarity, 60 ppm or less was more preferable. As described above, in the case of low-acid beverages such as milk tea that does not require much consideration of clarity and relatively strong taste, the addition concentration of the licorice organic solvent extract preparation can be increased, but barley tea can be used. In addition, the concentration of the addition was limited to a relatively low concentration in a beverage having a light taste such as

APPLICATION EXAMPLE 1 Barley tea raw material 80 g is converted to ion exchange water 1600 g at 90 ° C.
For 30 minutes, followed by filtration through filter paper (No. 2, Advantech Co., Ltd.) to remove the raw material residue and to extract 1440 g of barley tea extract (pH 4.9, Brix 0.6 °)
I got The barley tea extract was cooled to 30 ° C. or less, diluted with ion-exchanged water so as to have a drinking concentration (Brix 0.4 ° ), and sodium L-ascorbate and a licorice organic solvent extract preparation prepared in Example 3 With a final concentration of 300
ppm and 0.1% (30% as licorice organic solvent extract concentration)
ppm). Sodium bicarbonate was added thereto to adjust the pH to 6.0 to obtain a barley tea preparation. After filling this into a container, retort sterilization treatment (123
C. for 20 minutes), followed by cooling to room temperature with running water. The sterilized barley tea beverage is sterilized in advance with 5 ppm ozone water for 90 seconds, and then filled in a 2-liter PET bottle washed with sterilized water in a clean bench (class 100). It was sealed with the treated cap to obtain a barley tea beverage by aseptic filling.

Application Example 2 A mixed tea was trial-produced using a raw material containing 65% oolong tea, 20% black tea, 5% jasmine, 4% cinnamon, 4% hibiscus and 2% banaba. 30 g of the mixed tea was extracted with 900 g of ion-exchanged water at 90 ° C. for 10 minutes, and then filtered through a filter paper (No. 2, manufactured by Advantech Co., Ltd.) to remove the tea leaves and remove 820 g of the mixed tea extract (pH Four.
5, Brix 0.9 °, tannin concentration 150 mg / 100
mL).

The mixed tea extract was cooled to 30 ° C. or lower and diluted with ion-exchanged water to a drinking concentration (Brix 0.2 °). L-ascorbic acid and the licorice organic solvent extract preparation prepared in Example 3 were each brought to a final concentration of 200
ppm and 0.05% (15 ppm as the licorice organic solvent extract concentration), and sodium hydrogencarbonate was dissolved therein to obtain a mixed tea mixture having a pH of 6.0. This is filled in a container and subjected to a retort sterilization treatment (121 ° C., 15
Min), followed by showering with tap water until the temperature reached 30 ° C. This sterilized mixed tea beverage is
After sterilizing for 30 seconds at 40 ° C. using 0.15% P-3 oxonia active (manufactured by Ecolab, peracetic acid formulation), 500 mL PET bottles washed with sterilized water were filled in a clean bench (class 100). . The bottle was sealed with a cap that had been sterilized in the same manner as the bottle to obtain a mixed tea by aseptic filling.

Application Example 3 30 g of black tea was extracted with 900 g of ion-exchanged water at 70 ° C. for 5 minutes, and then filtered through a filter paper (No. 2, manufactured by Advantech Co., Ltd.) to remove tea leaves and remove 780 g of black tea. Extract (pH 5.5, Brix 1.1 °, tannin concentration 30
0 mg / 100 mL). 30 ℃ of the black tea extract
Cooled down to: The black tea extract, milk, sugar, emulsifier (DK Ester E-6000, Daiichi Kogyo Seiyaku Co., Ltd.)
The ethanol solution of 10% licorice organic solvent extract (2) and ion-exchanged water were 33: 25: 5: 0.2: 0.1: 36.
7 was mixed. To this was added an appropriate amount of sodium hydrogen carbonate to adjust the pH to 6.8 to obtain a milk tea preparation.

The mixture was stirred well while heating to 60 ° C. Thereafter, homogenization (homogeneous pressure: 200 kg / cm ² ) was performed using a homogenizer. This was filled in a container, and after retort sterilization treatment (121 ° C., 20 minutes), it was cooled to room temperature with water. Thereafter, a 1.5-liter PET bottle and a cap were sterilized in the same manner as in Application Example 1 , and then sterilized milk tea was filled and sealed in a clean bench to obtain aseptic-filled milk tea.

Application Example 4 Coffee extract (trade name: coffee extract M-0-20, Brix
20 ° , manufactured by Takasago Coffee Co., Ltd., milk, sugar, emulsifier (trade name: Sunsoft Super V-103, manufactured by Taiyo Chemical Co., Ltd.),
Each raw material of the licorice organic solvent extract preparation prepared in Example 3 and ion-exchanged water was 4.5: 10: 5: 0.2: 0.1: 80.
2 were mixed. An appropriate amount of sodium bicarbonate was added thereto to obtain a coffee beverage having a pH of 6.8. The mixture was stirred well while heating to 60 ° C. Thereafter, homogenization was performed with a homogenizer (homogenization pressure 200 kg /
cm ² ). This was filled in a container, and after retort sterilization treatment (121 ° C., 20 minutes), it was cooled to room temperature with water. Then, 500 m in the same manner as shown in Application Example 2
After sterilizing the L-volume PET bottle and the cap, the sterilized milk coffee was filled and sealed in a clean bench to obtain aseptic-filled milk coffee.

Application Example 5 30 g of green tea was extracted with 900 g of ion-exchanged water at 70 ° C. for 5 minutes, and then filtered through a filter paper (No. 2, manufactured by Advantech Co., Ltd.) to remove 800 g of green tea. Extract (pH 6.0, Brix 1.1 °, tannin concentration 270
mg / 100 mL). The green tea extract was cooled to 30 ° C. or lower, and the drinking concentration (tannin concentration 60 mg / 1
00 mL), and L-ascorbic acid and the licorice organic solvent extract preparation prepared in Example 3 were added to 200 ppm and 300 ppm, respectively, and sodium hydrogencarbonate was dissolved therein. PH
A green tea preparation of 6.0 was obtained. This was filled in a container, and subjected to a retort sterilization treatment (121 ° C., 7 minutes).
Cooled down. This was filled in a clean room (class 10,000) and sealed in a 500 mL PET bottle sterilized in the same manner as in Production Example 1 to obtain a green tea beverage.

[0055]

Industrial Applicability According to the present invention, it has been found that an organic solvent extract of a licorice or licorice water extraction residue has a remarkable growth inhibitory effect against sterilizing drug-resistant fungi, particularly against fungi belonging to the genus Earthrinium and Ketomium. . Such an antifungal effect of a licorice organic solvent extract has never been known before. In addition, the organic solvent extract of licorice or licorice water extraction residue is a natural product with a wealth of experience in foods, so it is highly safe and suitable for adding to non-heat-resistant packaged low-acid beverages. is there.

Therefore, if the antifungal agent of the present invention is added to a non-heat-resistant packaged low-acid beverage, particularly a non-heat-resistant packaged low-acid beverage produced via an aseptic filling and packaging system, serious problems may occur during the production of the beverage. The risk of contamination by the microorganisms, which is a serious problem, can be greatly reduced. In addition, the present invention can solve problems such as prolonging the sterilization time for sterilizing drug-resistant bacteria and using multiple sterilizing agents, thereby improving product quality and safety as well as improving productivity. It also has a remarkable effect on reducing the manufacturing cost.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification code FI A23L 2/44 A61P 31/10 3/3472 A23L 2/00 P A61P 31/10 W (58) Investigation field (Int.Cl. ⁷ , DB name) A61K 35/78 A23F 3/16 A23F 5/24 A23L 2/00 A23L 2/38 A23L 2/44 A23L 3/3472 BIOSIS (DIALOG) CA (STN) JICST file (JOIS)

Claims (3)

(57) [Claims] 1. A hypochlorite as a sterilizing agent, comprising an organic solvent extract of licorice or a licorice aqueous extraction residue.
Resistant to sodium citrate, hydrogen peroxide, peracetic acid, or ozone water
Antifungal agent effective for sexual fungi. 2. The antifungal agent according to claim 1 , wherein the drug-resistant fungus for sterilization is a fungus of the genus Arthrinium and / or Ketomium. 3. The antifungal agent according to claim 1, wherein the organic solvent extract of the licorice or licorice water extraction residue is obtained by extracting the licorice or licorice water extraction residue with ethanol or aqueous ethanol.