JPS6035104B2 - food preservatives - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention uses 300 to 500 PPM of lysozyme (W'V in the present invention).

) containing 50 to 75% ethanol (in the present invention, % simply means W'V).
) This relates to a food preservative that is characterized by having a remarkable effect on inhibiting the growth of ATCC6633, and the addition of a small amount of the product of the present invention inhibits the growth of microorganisms that cause spoilage without impairing the flavor of the food. The purpose is to provide a food preservative that inhibits food preservatives. In recent years, preservatives and preservatives are often added to foods distributed on the market, from raw materials to processed products, to maintain freshness, quality, and prevent spoilage.

However, on the other hand, the toxicity, carcinogenicity, and teratogenicity of these preservatives and preservatives have become problematic, and some substances have already been banned or restricted in use, and some are currently under consideration. As is well known.

It has been known that ethanol has the effect of killing microorganisms at high concentrations and suppressing their growth at low concentrations, and has already been used as a preservative in some foods.

However, if the amount of ethanol added is too large, the ethanol odor will become strong, impairing the flavor of the food, or denaturing the protein, and if too little is added, the desired effect may not be obtained. Due to the lack of natural ingredients, it was previously only used in foods with strong flavors such as sea urchin, miso, and soybean oil. However, the use of hydrogen peroxide has recently become a problem, and the bactericidal and microbial growth inhibiting effects of ethanol, which is considered harmless from a sanitary standpoint, have increased its reputation as a food preservative. This has led to the development of preservatives with formulations that do not alter the protein in food or impair flavor due to ethyl odor. “Neto” frequently occurs in foods containing ingredients such as
Bacillus subtilis (bag cucumber subtms) is the cause of so-called "rope" such as "rope" phenomenon.
) For ATCC6633, no preservatives with sufficient antiproliferative effects have been provided.

In the present invention, as a result of various studies, we have found that an ethanol aqueous solution in which a certain amount of lysozyme is dissolved not only more effectively suppresses the growth of microorganisms in foods than an ethanol aqueous solution of the same concentration that does not contain any lysozyme. , Bacillus subtilis (mother cillus mis) ATCC
The present invention was completed based on the discovery that the present invention has a particularly remarkable growth-inhibiting effect on 6633.

Since the lysozyme used in the present invention has the property of hydrolyzing bacterial cell membranes made of mucopolysaccharides, it is used as a powdered food preservative in the food industry either alone or in combination with other substances. There are some examples where lysozyme is used, but it is not very marketable, partly because lysozyme is expensive.

The ethanol aqueous solution containing lysozyme of the present invention is used in a liquid state, and by adding 3 to 4% to food and mixing it homogeneously, it can be used without impairing the flavor unique to the food. In addition, it effectively suppresses the growth of attached microorganisms without causing protein deterioration in food.
In particular, Bacillus subtilis (Bacillus subtilis)
lis) ATCC6633, and shows excellent results in preventing the occurrence of "neto" and "threading" phenomena, but in this case, the lysozyme contained in the product of the present invention The amount is preferably 300-500PP
M, since the ethanol concentration is 50-75%,
When the product of the present invention is added to food at a preferable concentration of 3 to 4%, the proportion of lysozyme to the food is 9 to 20%.
Although the ethanol content is low at 1.5-3.0%, the addition of lysozyme significantly enhances the growth-inhibiting effect of ethanol on microorganisms, especially Bacillus subtilis.
1 female subtilis) ATCC 6633 is not only seen in other food preservatives using lyzozyme, but also in food preservatives based on ethanol. It's something you don't see.

The lysozyme-containing aqueous ethanol solution according to the present invention is most preferably produced by dissolving 300 to 50 PM of lysozyme in an ethanol aqueous solution diluted to 58 to 75 vol% with purified water. It is preferable to add other organic acids such as lactic acid in order to maintain the optimum pH in connection with this, since organic acids themselves have the effect of killing and suppressing the growth of microorganisms. Next, a test example regarding the growth inhibiting effect of various spoilage bacteria of the lysozyme-containing ethanol aqueous solution according to the present invention will be explained.

Test Example 1 In Test Example 1, a bouillon pot with the following composition was used, and PseMomonasa (PseMomonasa), which is a typical bacterial strain related to food spoilage, was added to the broth.
Pseudomonas aeruginosa) LF05445 (hereinafter referred to as Pseudomonas aeruginosa).

), Proteus sp. (hereinafter referred to as Proteus bacteria), and Bacillus subtilis
ATCC 6633 (spore solution) (hereinafter referred to as Bacillus subtilis) was used for testing. The number of bacteria in the bacterial fluid during the action of these bacteria was 32 x 1 for Pseudomonas aeruginosa, 11 x 1 for Proteus on the skin, and 59 x 1 for Bacillus subtilis on the skin. {1} Normal broth medium (no additives) ■ Lysotium broth medium (medium made by adding 1 part of lysozyme 400 PPM aqueous solution to 28 parts of normal broth to make the lysozyme content 13.SPM) {3' Ethanol broth medium ( t4} Lysozyme-processed tanol/bouillon base (5 pieces containing 400 PPM of lysozyme in 28 parts of regular bouillon) To carry out Test Example 1, first, an L-shaped monoculture tube 4 was prepared.
Prepare 3 sets of books, collect 10 broth medium according to each of the above items into 4 culture tubes of each set, and then transfer to 4 culture tubes of the first set. One layer of Pseudomonas aeruginosa liquid was added to the second layer, one layer of Bacillus protuse liquid was added to the third layer, one layer of Bacillus subtilis spore solution was added to the third layer, and cultured by shaking at 370°C. Measure the light transmittance of the medium using a spectrophotometer every hour (every 3 minutes if necessary), and observe changes in transparency due to bacterial growth in Figure 1 for Pseudomonas aeruginosa and Figure 2 for Proteus bacteria. , and Bacillus subtilis were graphed and plotted as a pointillist diagram in Figure 3.

In Figure 1, the normal broth culture medium (1) and the lysozyme broth culture medium (■) begin to rapidly proliferate almost at the same time from around 4 hours, but there is a slight delay in the latter, indicating that bacteria are increasing in contact with the medium. The inhibition is thought to be due to the presence of lysozyme.

The results show that the ethanol/bouillon paste paste started to proliferate about 4 hours later, and had a stronger growth-inhibiting effect on bacteria than the lysozyme/bouillon paste, but the lysozyme-processed tanol/bouillon paste in [4] 3} more than 30
The fact that growth begins after a delay of about minutes and the time lag gradually widens means that when lysozyme is added to ethanol, it not only has a greater growth-inhibiting effect on bacteria, but also has a slight 10-Q effect. What is being expressed is observed. In Figure 2, the normal broth medium {1) and the lysozyme broth medium {2} both start to rapidly proliferate almost simultaneously from around 2.5 hours, but there is a slight delay in the latter. It is thought that the presence of lysozyme is responsible for the fact that bacterial growth is suppressed.

The ethanol/broth medium of [t3] and the lysozyme caroethanol/broth medium of [4] both started to grow bacteria after a further delay of about 30 minutes (indicating that they have a stronger growth-inhibiting effect on bacteria than the lysozyme/broth medium of 21). However, with the passage of time, the delay in growth time of [4L lysozyme-processed tanol/bouillon medium] became more and more pronounced than that of [3] ethanol/broth medium, and the growth-inhibiting effect on bacteria gradually became stronger. This is considered to be due to the presence of lysozyme.
The lysozyme broth solution {2} starts to grow about 1 hour later than that, but compared to the 30 minute delay for Pseudomonas aeruginosa in Figure 1 and the almost simultaneous growth start for Proteus in Figure 2, the growth of Bacillus subtilis is slower. The fact that the start time of growth is considerably delayed indicates that lysozyme has a stronger influence on Bacillus subtilis than on the other two bacteria. The ethanol/broth medium in ``3'' is about 3 times later in the start time of growth than the lysozyme/broth medium in ①, which suggests that the ethanol/broth medium has a stronger growth-inhibiting effect on bacteria than the lysozyme/broth medium. However, as time passes, the difference gradually decreases and eventually disappears. ■The lysozyme-added ethanol base is '
The growth of the bacteria started about 2 hours later than in the ethanol broth medium of 3), which is a significant difference from the aforementioned Pseudomonas aeruginosa and Proteus bacteria, which started growing at almost the same time. This indicates that ethanol added with lysozyme has a much stronger growth-inhibiting effect on Bacillus subtilis than on Pseudomonas aeruginosa and Proteus. Test Example 2 In Test Example 2, a bouillon pot with the following composition was used, and Proteus bacteria and Klebsieff pneumoniae were used as spoilage bacteria strains.
) ATCC9621 (hereinafter referred to as Klebsiella bacteria).

) and Bacillus subtilis (spore solution). The number of bacteria in the bacterial solution when these bacteria act is 35 x 107/' for Proteus bacteria and 12 x 1/' for Klebsiella bacteria.
and Bacillus subtilis 17×1.

m Ordinary bouillon base (no additives) ■ Ethanol bouillon base (medium containing 24 parts of regular bouillon and 1 part of a 75 vol% ethanol aqueous solution to make the ethanol content 3.0 vol%) '3' Lysozyme Processed tanol/broth medium (medium made by adding 1 part of a 75 vol% ethanol aqueous solution (product of the present invention) containing 400 PPM of lysozyme to 24 parts of ordinary broth to make 16 PPM of lysozyme and 3.0 mol% of lysozyme) of Test Example 2 The implementation method was carried out according to Test Example 1.

However, a bacterial solution of Proteus bacterium was added to the first set, a bacterial solution of Klebsiella was added to the second set, and a spore solution of Bacillus subtilis was added to the third set. 4 for Klebsiella, FIG. 5 for Bacillus subtilis, and FIG. 6 for Bacillus subtilis. In these Test Example 2, ordinary broth medium without additives was used as a control.
Ethanol 3. The trends in the growth inhibitory effects on Proteus bacteria, Klebsiella bacteria, and Bacillus subtilis were measured in the ethanol bouillon mixture containing 1% lysozyme and the lysozyme-treated tanol bouillon mixture to which lysozyme 1 bait PM was added. In the case of the ethanol broth medium (2), as shown in Figures 4, 5, and 6, the effect on all bacteria was almost the same, but
[In the case of 3 liters of lysozyme-processed tanol/fyon taji, the effect of adding lysozyme was added to the growth inhibition effect of eta/le bouillon taji against the first two bacteria. Of course, the fact that a synergistic growth-inhibiting effect is seen against Bacillus subtilis in addition to the latter effect is extremely characteristic, as seen in Figure 3 in Test Example 1. be.

Test Example 3 In Test Example 3, a bouillon pot with the following composition was used, and a spore solution of Bacillus subtilis was applied to test the trend of the effect of inhibiting bacterial growth.

In this case, the number of bacteria in the bacterial solution during the action of Bacillus subtilis is 1
It was 7×1 pi/pan'.

{1} Ordinary broth medium (no additives) ■ Ethanol bouillon toji (Nagisaji, where an aqueous solution of 75 vol% ethanol was added to 24 parts of ordinary bouillon to bring the ethanol content to 3.0 vol%) '3' bottles Bouillon base with addition of invention product [1 part of the invention product (75 vol% ethanol aqueous solution containing 400 PPM of lysozyme and 0.2% lactic acid) was added to 24 parts of ordinary broth medium to reduce the ethanol content to 3. 4) Broth culture medium with product A added [24 parts of ordinary bouillon base and 1 part of company A's product (75 vol% ethanol aqueous solution containing 2.2% special brewed vinegar and 0.5% lactic acid) 3.5 parts to bring the ethanol content to 3. '5' Broth medium with 8 products added [24 parts of ordinary bouillon base and 1 part of Company B's product (75 vol% ethanol aqueous solution containing 0.1% malic acid and 0.4% glycine) was added to reduce the ethanol content to 3. Medium and ol%] Test Example 3 was carried out in accordance with the previous two tests, and the progress was graphed in FIG. 7.

Test Example 3 was a comparative test of the trend in the bacterial growth inhibitory effect on Bacillus subtilis (spore solution) between a product made by another company mainly containing ethanol and the product of the present invention. Set the joint rate to 3. The same concentration was set as Wol%.

In this test, the normal broth medium in [1] grew seriously from around the 4th hour, and rapidly increased from around the 5th hour, whereas the ethanol broth medium in '2), ■
There is a slight time difference in the order of the A product-added broth medium and [5'B product-added broth medium: '2}, '41, t5}, but growth begins almost simultaneously around the 6th hour, and the 7th hour begins. Since then, the growth has been rapid, and the time difference between the growth of the bacteria in the ordinary broth medium and '1' is about 2 hours. On the other hand, in the bouillon solution containing the present invention product added at t3}, proliferation started at around 8 hours and increased rapidly from around 9 hours. }, [4' ethanol bouillon beach area,
The fact that the growth start time is about 2 hours later than in the broth medium etc. supplemented with products A and B indicates that the inhibitory effect on bacterial growth is that strong.Simply put, In addition to its general preservative effect, the product of the present invention has a growth-inhibiting effect against Bacillus subtilis that is approximately twice as effective as preservatives containing glycine or other key organic acids as one of its active ingredients. One thing can be said.

[Brief explanation of the drawing]

Figure 1 shows PseMomonasaem bonito nosa IF054.
45, Figure 5 shows Klebsiyev Neumonia (K1ebsj
e11apneumoniae) ATCC9621, Figures 2 and 4 are f. Proteussp, Figures 3, 6 and 7 are Bacillus subtilis (European cmussubtili).
s) The growth trend of each bacteria of ATCC6633 (spore liquid) on various media was measured over time by absorbance (wavelength: 54 m).
). Figure I Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7

Claims (1)

[Claims] 1. Bacillus subtilis mainly in a 50-75% (by weight) aqueous solution of ethanol containing 300-500 PPM (by weight) of lysozyme.
lis) A food preservative that is highly effective in inhibiting the growth of ATCC6633.