JPH05252928A - Method for preventing generation of mold on leaf tobacco - Google Patents

Abstract

PURPOSE:To economically prevent the generation of mold on leaf tobacco during its storage without affecting the flavor and smoking taste of the leaf tobacco by harvesting the leaf tobacco, and subsequently inoculating a Bacillus bacterium on the surface of the leaf tobacco arbitrary times at an arbitrary time. CONSTITUTION:Leaf tobacco is harvested and subsequently inoculated with a bacterium belonging to the Bacillus (preferably Bacillus.amyloliquefaciens) arbitrary times at an arbitrary time until the leaf tobacco is dried with air, thus performing the objective mold generation-preventing treatment.

Description

Detailed Description of the Invention

[0001]

FIELD OF THE INVENTION The present invention relates to a method for preventing mold growth in leaf tobacco of air-dried seeds.

[0002]

2. Description of the Related Art Conventionally, there has been a problem in that mold development during drying or storage of air-dried seed tobacco. This mold is
It occurs because molds belonging to the genus Aspergillus, the genus Penicillium, the genus Alternaria, etc. parasitize leaf tobacco of air-dried species. Tobacco, which has a high degree of mold development, has a poor aroma and flavor, resulting in a decrease in commercial value. Moreover, the significantly deteriorated cigarettes have to be discarded, resulting in a large annual loss. Therefore, a great deal of labor and cost is currently spent on the prevention of mold.

Generally, the types of cigarettes are roughly classified into
It is divided into three types, Burley, Native and Yellow. The leaf tobacco is dried after harvesting from the field, and the drying method depends on the type of tobacco. Forced drying is performed on yellow species, while natural drying in air is performed on Burley and conventional species, ie air dried species.

Air-drying is carried out for about 45 days, and the drying period is roughly classified into a yellowing period, a browning period and a mid-bone drying period. When drying air-dried tobacco, it is necessary to maintain appropriate humidity conditions during these entire periods in order to prevent rapid dehydration. Therefore, since the humidity is maintained to some extent, it is likely to grow. The browning period starts from about the 15th day, and the cells die during this browning period, so that molds are particularly likely to grow. Also, in the middle part of the leaf tobacco, the water does not evaporate to the end, so the mold grows more easily than other parts.

On the other hand, dried leaf tobacco is stored in a warehouse of a cultivating farm or a raw material factory, but it may absorb moisture during the storage period to form a mold.

[0006] At present, as a method for preventing the growth of mold in the case of natural drying, the relative humidity is decreased by heating in the drying process, the storage bag is used for preventing moisture absorption in the storage process, and especially in the case of cultivated farmers. Brushes are used to remove mold on the surface of leaf tobacco, and leaf tobacco is regularly reloaded in factories. However, there is a limit to lowering the relative humidity by heating. In addition, in the case of removal with a brush or transshipment, the work is heavy labor,
Moreover, since it is performed by human hands, there is a problem that allergic symptoms occur during the removal work.

[0007] Further, conventionally, in the drying process, spraying of a bactericide or use of a fumigant has also been examined, but none of them can be used because of adverse effects on the flavor and taste of tobacco and safety problems such as pesticide residue. It was

[0008]

SUMMARY OF THE INVENTION Therefore, an object of the present invention is to provide a method for preventing mold growth of leaf tobacco which has no adverse effect on the flavor and taste of tobacco and is effective in terms of economy and workability. ..

[0009]

[Means for Solving the Problems] In order to solve the above problems, the present inventors have used an antifungal microorganism that grows on leaf tobacco when sprayed on the leaf tobacco after harvesting until the end of drying treatment. In order to study the antifungal method, various microorganisms were isolated from the leaf tobacco drying room, soil, and the developed leaf tobacco, and intensive research was conducted on their antifungal and colonization properties. As a result, they have found that bacteria belonging to the genus Bacillus exhibit a high antifungal effect, and have completed the present invention based on this finding.

That is, according to the present invention, when leaf tobacco is harvested, dried in an air drying step, and the dried leaf tobacco is stored until use, it is added to the genus Bacillus at any time after harvesting. Provided is a method for preventing mold growth of leaf tobacco, which comprises inoculating the surface of a leaf tobacco with a bacterium belonging thereto any number of times.

The method of the present invention is applied to tobacco dried in an air drying process. That is, typically
It is applicable to any type of tobacco, including but not limited to Burley and native species, as long as it is air dried.

The present invention will be described in detail below.

First, the bacteria used in the method for preventing mold growth of the present invention are cultured.

As the bacterium used in the method for preventing fungal growth of the present invention, any bacterium belonging to the genus Bacillus can be used, but Bacillus amyloliquefaciens has a particularly high antifungal effect. preferable.

Bacillus amyloliquefaciens (Ba
cillus amyloliquefaciens) examined the mycological properties of this bacterium according to the method described in the Manual of Microbiological Method, and based on this, the Burgess Manual of Bergey's Manual of Determinative Bacteriolog
y 8th Edition) and can be identified by searching.

The bacteriological properties of this bacterium are shown below.

I. Morphological properties 1. Microscopic findings Bacillus. 0.9 × 4.0 to 4.5 μm, wavy linkage, elliptic endospore formation, 1 per cell, central position, gram positive, acid-negative. 2. Cultivation findings (1) Plate culture Irregularity, flat upper surface, fine granules, surface reticulate, serrated edge, gray. (2) Slope culture It has a vigorous growth, is smooth, spreads over the entire surface, and has a serrated, slightly gray edge. Slight ammonia smell. (3) Medium growth, white film is formed on the surface. Little turbidity.
Little precipitation.

II. Physiological properties (1) Growth temperature: 15 to 42 ° C, optimal 30 to 37 ° C (2) Growth pH: pH 3.5 to 10.0, optimal 6.5
7.5 (3) Oxygen requirement: aerobic (4) Liquefaction of gelatin: Liquefied in a funnel shape (5) Litmus milk: acidic, peptone (6) Indole formation: negative (7) Reduction of nitrate: Positive (8) Starch hydrolysis: Positive (9) Methyl red reaction: Negative (10) Forges Broschauer reaction: Positive (11) Catalase production: Positive (12) Urease production: Negative (13) Salt concentration Growth: Growth up to 7% (14) Growth inhibition by sodium azide: 0.02
% Growth (15) Growth inhibition by lysozyme: 0.001% growth (16) Sabouraud agar medium growth: Grow (17) Sabouraud liquid medium growth: Grow (18) 0-F test : Fermentative (19) Oxidase activity: positive (20) Utilization of citric acid: positive (21) Lecithinase activity: negative (22) Tyrosinase activity: negative (23) Levan production: positive (24) Casein hydrolysis: positive (25) Deamination reaction of phenylalanine: Negative (26) Generates acid and does not generate gas: glucose,
Arabinose, xylose, mannitol The Bacillus amyloliquefaciens that can be used in the present invention can be obtained from the Fermentation Research Institute (deposit number IFO14141).

As the medium for culturing the bacteria used in the present invention, a broth medium known for bacterial culture can be used. However, the present invention is not limited to this, and any medium may be used as long as it is used as a normal bacterial medium.

The culture conditions for culturing the bacterium used in the present invention include a temperature of 30 to 42 ° C. and a pH of 6.0 to 8.0.
It is preferably in the range of. More preferably temperature 37
C., pH 7.0. The bacteria used in the present invention are aerobic and thus require oxygen.

Since the bacterium used in the present invention forms spores in the latter half of the culture, it is preferable to terminate the culture at the time when a large number of spores have been formed after the logarithmic growth phase and used. The bacterium used in the present invention exhibits an antifungal effect in the state of vegetative cells after germination of spores. When a large amount of spores are formed, a large amount of spores are present in addition to the vegetative cells, and these spores also germinate to become vegetative cells and exert their effects. Therefore, it is most effective from a quantitative point of view to use it at this point.

After culturing the bacterium of the present invention for a certain period of time, the bacterium is collected by centrifugation. The collected bacterial cells are suspended in a specific buffer solution to prepare a bacterial solution.

As the buffer for suspending the cells of the present invention, it is preferable to use water or Seleensen's phosphate buffer.

The ratio of the present cells suspended in a buffer solution is as follows:
It is preferable to suspend 10 ⁹ to 10 ¹⁰ cells per 1 ml of the buffer solution.

When this bacterium solution is not used immediately for the mildew-proof treatment, it is desirable to store it at a low temperature in order to prevent contamination of various bacteria. The storage temperature is preferably 1 to 5 ° C.

Next, the fungus body obtained by the above-mentioned method is subjected to a mildew-proofing treatment for leaf tobacco.

First, water is added to the above-mentioned bacterial solution containing the required amount of bacteria to prepare an inoculum, and this inoculum is sprayed uniformly on leaf tobacco to inoculate. This bacterial species may be used immediately after harvesting and at any time during storage.

The preferred inoculation time is immediately after harvesting or during the browning stage of the air drying process, but immediately before entering the drying process, the yellowing period or mid bone drying stage of the air drying process, immediately after the completion of the drying process,
It may be carried out at an appropriate time during storage.

The surface area (total of front and back) per leaf tobacco used in the present invention is 400 in the case of the conventional species.
~ 2000 cm ² , 500-250 for Burley
It is 0 cm ² .

The amount of inoculum is 10 ⁵ to 1 leaf tobacco
It is 10 ⁸ pieces, preferably 10 ⁷ to 10 ⁸ pieces, and the amount of inoculum required for spraying is 2-3 ml per leaf for leaf tobacco immediately after harvesting and in the early stage of drying, and 0.5 for leaflets after the middle stage of drying. ~ 1 ml. Therefore, 20 cells
It is preferable to prepare an inoculum by suspending in 50 times the amount of water.

By spreading the spray solution thus prepared on the target tobacco leaf, it is possible to effectively prevent the growth of mold.

[0032]

EXAMPLES Examples of the present invention will be described below.

In Examples 1 and 2, Burley seeds having a surface area of 1500 to 1800 cm ² per leaf tobacco were used.

Example 1 Bacillus was added to a broth medium (meat extract broth: meat extract 10 g, peptone 10 g, salt 1 g, distilled water 1 liter).
Inoculated with Amyloliquefaciens IFO14141,
The cells were cultured at 37 ° C for 72 hours with shaking. The culture medium containing the bacterial cells was centrifuged at 5,000 rpm to separate the bacterial cells, and the cells were suspended in a phosphate buffer of Seelensen and washed. The cells were collected by centrifuging again and resuspended in 40 times the amount of the culture solution of phosphate buffer of Selezenen to obtain 10 ⁹ cell solution per ml.

To 2 ml of the bacterial solution obtained by the above method, 48 ml of tap water was added and diluted 25 times to prepare an inoculum of 50 ml. As a treatment group, this inoculum was sprayed on 50 pieces of dried leaf tobacco. That is, it means that 4 × 10 ⁷ Bacillus amyloliquefaciens was applied to each leaf tobacco.

As a control, 50 ml of tap water was sprayed on 50 leaf tobaccos.

After both sections were left at 25 ° C. for 24 hours, the water content was adjusted to 15% (dry base) and placed in a constant temperature and constant humidity box adjusted to a temperature of 25 ° C. and a humidity of 85 to 95%. Aspergil
Lusochraceus and Penicillium species were slant-cultured in a test tube to form conidia, and spores of one test tube (about 10 ⁵ as the spore amount) were sprayed on each section.

This was stored for 2 months and the mold development status of leaf tobacco was compared in both plots. The effect of the treatment of the present invention on the mildew is shown in Table 1 below.

[0039]

[Table 1] Some mold was found along the middle bones in part of the treatment area,
Compared with the control group, the mold-inhibiting effect is high, and it can be judged that the mold-inhibiting effect by this treatment is high.

Furthermore, the effects of this treatment on the flavor and taste of tobacco were compared between the two plots. The treated section and the control section were wound up on a hand-rolled cigarette, and a trial was carried out using an in-house panel. 1:
The results of discriminating the same products by the 2-point method are shown in Table 2 below.

[0041]

[Table 2] From Table 2 above, it can be said that even if the leaf tobacco is subjected to the treatment of the present invention after the Burley seeds are dried, it has no effect on the flavor and taste of the tobacco.

Example 2 A bacterial solution obtained by the same method as in Example 1 was diluted 20 times with tap water to prepare an inoculum of 5 × 10 ⁷ cells / ml.

Immediately after harvesting, 100 sheets of Burley leaf tobacco were inserted in a petiole of the hemp rope and continuously knitted so that the leaves were back-to-back and back-to-back.

As a test section, this inoculum was sprayed onto the above-mentioned continuous weave leaf tobacco in an amount of 2 ml per sheet.
That is, it means that 10 ⁸ pieces were scattered per sheet.
As a control, 2 ml of tap water was sprayed per leaf tobacco.

Both of the sections were suspended in a pipe house for 14 days for yellowing, and then suspended in a wooden drying chamber for browning and further mid-bone drying for a total process of 45 days. On the 30th day after the start of the main drying treatment, a mold development survey was conducted.

Further, after the completion of the drying treatment, leaf tobacco (water content: about 12% (dry base)) was stored in a thermo-hygrostat box adjusted to 25 ° C. and 85-95%, and one month later, a mold development test was conducted. .. Table 3 below shows the results of the two molds investigations.

[0047]

[Table 3] From Table 3 above, it can be seen that by performing the treatment immediately after harvesting, mold development during the drying period and during storage can be prevented.

[0048]

EFFECTS OF THE INVENTION By using the method for preventing mold growth of the present invention, first, in a cultivated farmer, there is no need for the mold growth removal work and the amount of waste of the mold product can be reduced, so that the production amount is reduced. Can be prevented. In the raw material factory, it is possible to reduce the number of times of mold prevention control during storage in the warehouse, and to reduce the amount of defective raw materials discarded due to mold, so that the manufacturing cost can be reduced. Furthermore, since allergic symptoms do not occur during the mold removal work, improvement of human health can be expected.

 ─────────────────────────────────────────────────── ─── Continuation of the front page (72) Inventor Junichi Miki 1900 Idei, Oyama City, Tochigi Prefecture Japan Tobacco Inc. Leaf Tobacco Research Institute

Claims (2)

[Claims] 1. When harvesting leaf tobacco, drying it in an air drying process, and storing the dried leaf tobacco until use,
A method for preventing mold growth of leaf tobacco, which comprises inoculating the surface of a leaf tobacco with a bacterium belonging to the genus Bacillus at an arbitrary time after harvesting at an arbitrary number of times. 2. A bacterium belonging to the genus Bacillus is Bacillus amyloliquefaciens.
The method for preventing mold growth of leaf tobacco according to claim 1, which is a loliquefaciens).