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Method for separating microorganisms on surface of tobacco leaf at high temperature by utilizing pure tobacco leaf leach liquor solid plate

A solid plate and extraction liquid technology, applied in the field of applied microorganisms, can solve the problems of single action point, poor temperature tolerance, uneconomical economy, etc., and achieve the effect of improving physiological activity, reliable safety, and high biological activity.

Inactive Publication Date: 2011-06-01
KUNMING GUIXUN TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Although experiments have shown that the use of enzyme preparations can improve the quality of cured tobacco leaves, and has the advantages of single action point and easy control of action concentration, there are still some problems in the use of enzyme preparations during the curing stage.
First of all, most of the commercial enzymes currently on the market are room temperature enzymes, which have poor temperature tolerance and are not suitable for application under baking conditions.
Secondly, the general market price of high-temperature enzymes is relatively expensive, and the application cost is too high, which is not economically cost-effective.
Furthermore, in essence, enzymes are also a kind of protein. It is unreasonable to add protein to the upper tobacco leaves, which are already high in protein content.

Method used

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  • Method for separating microorganisms on surface of tobacco leaf at high temperature by utilizing pure tobacco leaf leach liquor solid plate
  • Method for separating microorganisms on surface of tobacco leaf at high temperature by utilizing pure tobacco leaf leach liquor solid plate
  • Method for separating microorganisms on surface of tobacco leaf at high temperature by utilizing pure tobacco leaf leach liquor solid plate

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1: Fermentation process parameters of bacterial strain YN91 and bacterial strains YN114, YN113, YN104 of the present invention. The fermentation medium of strain YN91 is: wheat bran 1.5%, urea 0.5%, NaCl 0.5%. Fermentation process parameters: temperature 50°C, time 18h, stirring speed 125rpm. The enzyme activity level is around 6960U / mL. The fermentation medium of strain YN114 is: soluble starch 0.5%, beef extract 1.0%, CaCO 3 0.3%. Fermentation process parameters: temperature 43°C, time 36h, stirring speed 125rpm. The enzyme activity level is around 28(U / mL). The fermentation parameters of strains YN113 and YN104 were the same as those of YN114.

[0033] The specific method is as follows: culture medium preparation, take 5-15mL of the waste tobacco leaf extraction stock solution soaked for 2-3 days in a 500mL triangular flask, add 95-85mL of tap water, add 2.5g of agar powder, the pH is natural, after shaking well, use 15 lb. Autoclave for 30 minutes. Af...

Embodiment 2

[0037] Example 2: The high-temperature bacteria strain YN114 isolated by the method of the present invention forms milky white colonies on the solid medium, which are opaque, with irregular edges, slightly raised center, wrinkled texture, and are usually relatively dry. The average length of the bacteria is about 3.3 μm, and the average width is about 1.4 μm. The electron microscope photos are as follows: figure 1 . The optimum growth temperature is 50°C, protease is produced, and the protease activity of the fermentation broth reaches 28.25U / mL.

Embodiment 3

[0038] Example 3: The bacterial strain YN91 isolated by the method of the present invention forms a milky white colony on a solid medium, which is irregular, opaque, wrinkled in texture, and wavy at the edge. The shape of the colony changes greatly with the dryness and humidity of the medium. When it is wet, the colony is easy to spread and becomes slimy, and the surface is creamy. However, in most cases, the colony is rough and wrinkled, with a prominent center and irregular edges. The bacteria grow rapidly. Observed under the oil microscope, YN91 is transparent and shiny, short rod-shaped, does not produce spores, 2.6μm-1.1μm, see the electron microscope photo figure 2 . Strain YN91 belongs to Pseudomonas pridus. Produce amylase, the optimum growth temperature is 55°C for 30 hours, and the enzyme activity of the fermentation broth is 7840U / mL.

[0039] The present invention adds 0.1%-0.5% MnSO to the fermentation liquid before the end of fermentation 4 After stirring for...

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Abstract

The invention provides a method for separating microorganisms on a surface of a tobacco leaf at a high temperature by utilizing a pure tobacco leaf leach liquor solid plate, belonging to the technical field of applied microbiology. The technical scheme of the method comprises the following steps: mixing a waste tobacco leaf liquid which is fully soaked (for 3-5 days) with agar (2.5g / 100ml) based on the volume ratio of 5-15% (V / V), autoclaving at the pressure of 0.11-0.13MPa for 25-35 minutes (sterilizing for 230 minutes according to the ratio of 15 pounds per inch) under the conditions that no additional component is added and the pH value is not regulated, and pouring the mixture into a double layer plate to prepare the pure tobacco leaf leach liquor solid plate; and carrying out serial dilution on a tobacco leaf sample according to the conventional process, adding the diluted tobacco leaf sample into the pure tobacco leaf leach liquor solid plate, uniformly coating, and cultivating the pure tobacco leaf leach liquor solid plate in an incubator at the temperature of 50-60 DEG C until a bacterial colony is produced. The quantity and type of microorganisms grown on the surface of the tobacco leaf at a high temperature can be really reflected, and the microorganisms separated by the method are applicable to improving the quality of the tobacco leaf when the tobacco leaf is cured by flue.

Description

technical field [0001] The invention provides a method for high-temperature separation of microorganisms on the surface of tobacco leaves with pure tobacco leaf extract liquid on a solid plate, and belongs to the technical field of applied microorganisms. Background technique [0002] Microbial isolation method is an important technique to obtain pure cultured microorganisms. Microorganism isolation methods usually include medium composition and isolation culture temperature. According to the content of coagulant, the medium can be divided into solid, semi-solid and liquid medium. The separation temperature is also divided into low temperature, medium temperature and high temperature according to the cultivation temperature. With the development of the tobacco industry, microbial technology has been widely used in the tobacco industry. Microbial application technology has penetrated into the pre-production process of the cigarette industry, such as the tobacco agricultura...

Claims

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Application Information

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IPC IPC(8): A24B15/20A24B15/24C12N1/20
Inventor 杨程潘杰王毅赖永红张家伟
Owner KUNMING GUIXUN TECH
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