CN107384756B - Device and method for collecting tobacco essence and spice polluted microorganisms - Google Patents

Abstract

The invention relates to a device for collecting microorganisms polluted by tobacco essence and spice. Comprising an upper half shell and a lower half shell which can be combined together; wherein the upper half shell has a sample inlet and a sealing cover; the lower half shell is provided with a filter membrane and a bracket which can be combined. The collection device for the tobacco essence and spice polluted microorganisms has the advantages of simple structure, convenient operation and low cost, can effectively remove the influence of various essence and spice components on subsequent microorganism analysis, and solves the problem of cell loss caused by repeated sample transfer and operation steps in the microorganism collection process; meanwhile, the device can be integrally sterilized under high pressure, and is small in size and convenient to operate in an ultra-clean workbench, so that the risk of exogenous microorganism pollution is greatly reduced.

Description

A device and method for collecting microorganisms contaminated by tobacco flavors and fragrances

Technical field

The present invention relates to the technical field of collecting microbial cells, specifically a collection device for collecting microorganisms contaminated by tobacco flavors and fragrances.

Background technique

Microbial contamination is prevalent in all aspects of production, processing, transportation, and storage of food, drugs, and cosmetics, and is one of the important hidden dangers affecting product quality and safety. High-throughput sequencing technology overcomes the shortcomings of traditional culture, greatly deepens people's understanding of the structure and diversity of microbial flora in these samples, and promotes the effectiveness of prevention and control measures. Among them, the collection of microorganisms in samples is the first and very critical link in high-throughput sequencing technology, which directly affects the reliability of bacterial community structure and diversity analysis results. At present, the collection of contaminating microorganisms in these samples mainly adopts a combination of centrifugation, membrane filtration, oscillation and dispersion technology. However, there are many steps in the operation, which requires the use of filtration, washing, oscillation and other instruments, and the use of pipettes multiple times to collect microorganisms. Transferring will cause losses again.

Tobacco flavors and fragrances are indispensable and important raw materials for cigarette production. They can give cigarette products a unique style, improve smoke quality, and enhance product acceptability. They are a mixture of various spices, appropriate amounts of solvents, and other ingredients. In the production, storage and transportation of tobacco flavors and fragrances, quality degradation caused by microbial contamination leads to product scrapping. High-throughput sequencing technology is conducive to a more comprehensive understanding of the microorganisms that may exist in flavors and fragrances, providing prior risk warning for the prevention and control of spoilage of flavors and fragrances and formulating effective prevention and control measures to ensure product quality and safety. Compared with other types of samples, the components of tobacco flavors and fragrances are more complex and diverse. The numerous operating steps and multiple transfers of existing microbial collection methods inevitably lead to losses and affect the understanding of contaminating microorganisms. In addition, the microbial collection process Certain components remaining in the product will affect subsequent PCR amplification reactions.

Contents of the invention

In order to solve the above technical problems, the purpose of the present invention is to provide a collection device for microorganisms contaminated by tobacco flavors and fragrances, so as to remove the influence of various flavors and fragrances components on subsequent microbial analysis and the repeated transfer of samples and operating steps during the microbial collection process. Many problems cause cell loss; at the same time, the device can be fully autoclaved, and its small size makes it easy to operate in a clean workbench, thus greatly reducing the risk of contamination by external microorganisms.

The technical solution of the present invention is as follows:

A collection device for collecting microorganisms contaminated by tobacco flavors and fragrances is composed of: an upper half shell and a lower half shell that can be combined together; wherein the upper half shell has a sample inlet and a sealing cover; the lower half shell The outer shell has a filter membrane and a holder that can be combined.

The upper half shell and the lower half shell are both semi-elliptical cavities.

The upper half shell and the lower half shell are connected through threads on the inner wall of the shell.

The filter paper and the outer bottom of the filter membrane are snap-connected to the bracket, and the bracket is double-layered.

The number of each layer of the brackets is no less than 3.

The filter paper is close to the mouth of the lower half of the housing, and has a pore diameter of 35-45 μm.

The filter membrane is located near the bottom of the lower half of the housing and has a pore size of 0.22-0.45µm.

30-50mg glass beads can be placed in the lower half of the shell as needed.

Methods for the collection device to collect microorganisms contaminated with tobacco flavors and fragrances:

a. Connect the upper half of the shell to the lower half of the shell without the filter membrane and bracket, open the sealing cover of the upper half of the shell, put the glass beads and the sample to be measured in sequence from the sample inlet, and close the sealing cover. , shake thoroughly to disperse the sample, the shaking speed is 140-180rpm, and the shaking time is 10-15min;

b. Place the upper half of the shell below so that the sample is inside the upper half of the shell. Separate the lower half of the shell and connect the upper half of the shell containing the sample to the lower half of the shell containing the filter paper, filter membrane and bracket. Connect, where the pore size of the filter paper near the mouth of the lower half of the shell is 35-45μm, and the pore size of the filter membrane near the bottom is 0.22-0.45µm. The upper half of the shell is placed above, so that the sample passes through the filter paper and filter membrane sequentially;

c. Separate the upper and lower shells, take out the filter paper near the mouth of the lower shell in turn, connect the upper and lower shells, and add sterile deionizer from the sample inlet. 10-20mL of ionized water, the collected microorganisms are on the filter membrane for subsequent microbial analysis.

The invention has a simple structure, is easy to operate, and is low in cost. It can effectively remove the influence of various flavors and fragrance components on subsequent microbial analysis, and solves the problem of repeated sample transfer and multi-step operation steps causing loss to cells during the microbial collection process; at the same time, the invention The device can be autoclaved as a whole, and its small size makes it easy to operate in a clean workbench, thus greatly reducing the risk of contamination by external microorganisms.

Description of the drawings

Figure 1 is a schematic diagram of the overall structure of the present invention.

Figure 2 is a schematic structural diagram of the lower half of the casing of the present invention.

The reference symbols have the following meanings:

1. Upper shell; 2. Lower shell; 3. Thread; 4. Sample inlet; 5. Sealing cover; 6. Glass beads; 7. Filter paper; 8. Filter membrane; 9. Bracket.

Detailed ways

As shown in Figures 1 and 2, the device of the present invention is composed of an upper half shell 1 and a lower half shell 2 that can be combined together. The upper half shell 1 has a sample inlet 4 and a sealing cover 5. There are two types of lower shells 2. One has filter paper 7, filter membrane 8 and a bracket 9 that can be combined with them. The upper shell 1 and the lower shell 2 are both semi-elliptical cavities. , connected through the thread 3 on the inner wall of the shell; the bracket in the lower half of the shell 2 is double-layered, and the number of brackets in each layer is not less than 3; the outer bottom of the filter paper 7 and the filter membrane 8 are snap-connected to the bracket 9; the filter paper 7 Close to the mouth of the lower shell 2, the pore diameter is 35-45 μm, and the filter membrane 8 is close to the bottom of the lower shell 2, with a pore diameter of 0.22-0.45 μm; 30-30 μm can be placed in the lower shell 2 as needed. 50mg glass beads 6.

During use, first connect the upper half of the shell 1 to the lower half of the shell 2 without the filter membrane and bracket, open the sealing cover 5 of the upper half of the shell 1, and put in the glass beads 6 and the waiting liquid in sequence from the sample inlet 4. Measure the sample, cover it with the sealing cap 5, and shake it thoroughly to disperse the sample.

Put the sample in the upper half of the housing 1 below, separate the lower half of the housing 2, and connect the upper half of the housing 1 containing the sample and the lower half of the housing 2 containing the filter membrane and bracket. The upper half of the housing 1 is connected and the sample is sequentially passed through the double-layer filter membranes of 30-50μm and 0.22-0.45μm.

Separate the upper shell 1 and the lower shell 2, and take out the filter membranes near the mouth and bottom of the lower shell 2 in sequence. The collected microorganisms are the filter membranes near the bottom for subsequent microbial analysis.

Application Example 1:

During use, first connect the upper shell 1 to the lower shell 2 that does not contain filter paper, filter membrane and bracket, open the sealing cover 5 of the upper shell 1, and put glass beads 6 in sequence from the sample inlet 4 and the sample to be tested, cover it with the sealing cover 5, and shake it thoroughly to disperse the sample evenly, with a shaking speed of 140 rpm and a shaking time of 15 minutes;

Then place the upper half of the shell 1 below so that the sample is in the upper half of the shell 1, separate the lower half of the shell 2, and connect the upper half of the shell 1 containing the sample with the filter paper 7, filter membrane 8 and The lower half of the housing 2 of the bracket 9 is connected, and the upper half of the housing 1 is placed above, so that the sample passes through the filter paper and filter membrane in sequence;

Then separate the upper half of the shell 1 and the lower half of the shell 2, take out the filter paper 7 near the mouth of the lower half of the shell 2, and then connect the upper half of the shell 1 and the lower half of the shell 2, from the sample inlet 4. Add 10-20 mL of sterile deionized water, and the collected microorganisms will be on the filter membrane 8 for subsequent microbial analysis.

Application Example 2:

The sample dispersion and oscillation speed in this example is 160 rpm, and the oscillation time is 12 minutes. Other procedures are the same as Application Example 1.

Application Example 3:

The sample dispersion and oscillation speed in this example is 1180 rpm, and the oscillation time is 10 minutes. Other procedures are the same as Application Example 1.

Claims (5)

1. A method for collecting microorganisms contaminated with tobacco flavors and fragrances, characterized in that it is based on a microorganism collection device, which includes an upper half shell (1) and a lower half shell that can be combined together (2); wherein the upper half of the housing (1) has a sample inlet (4) and a sealing cover (5); the lower half of the housing (2) contains a filter paper (7) and a filter membrane (8) , the filter paper (7) and the filter membrane (8) are supported by the bracket (9), and the upper half shell (1) and the lower half shell (2) are both semi-elliptical cavities; The method for collecting microorganisms contaminating tobacco flavors and fragrances includes the following steps: a. Connect the upper shell (1) to the lower shell (2) without filter paper (7), filter membrane (8) and bracket (9), and open the upper shell (1). Seal the cover (5), put the glass beads (6) and the sample to be measured sequentially from the sample inlet (4), cover the sealing cover (5), and oscillate fully to disperse the sample. The oscillation speed is 140-180 rpm, and the oscillation time is 10-15 minutes. ; b. Place the upper half of the shell (1) below so that the sample is inside the upper half of the shell (1). Separate the lower half of the shell (2) and connect the upper half of the shell (1) with the sample. The lower half of the shell (2) containing the filter paper (7), the filter membrane (8) and the bracket (9) is connected, and the filter paper (7) near the mouth of the lower half of the shell (2) has a pore diameter of 35-45 μm. The filter membrane (8) near the bottom has a pore size of 0.22-0.45µm, and the upper half of the shell (1) is placed above, so that the sample passes through the filter paper (7) and the filter membrane (8) sequentially; c. Separate the upper half of the shell (1) and the lower half of the shell (2), take out the filter paper (7) near the mouth of the lower half of the shell (2), and then remove the upper half of the shell (1) Connect to the lower half of the shell (2), add 10-20mL of sterile deionized water from the sample inlet (4), and the collected microorganisms will be on the filter membrane (8) for subsequent microbial analysis. 2. A method for collecting microorganisms contaminated with cigarette flavors and fragrances according to claim 1, characterized in that there is a shell between the upper half shell (1) and the lower half shell (2). Threaded (3) connection to the body wall. 3. A method for collecting microorganisms contaminated by tobacco flavors and fragrances according to claim 1, characterized in that the outer bottom of the filter paper (7) and the filter membrane (8) are engaged and connected with the bracket (9), and the bracket (9) 9) It is double layer. 4. A method for collecting microorganisms contaminated with tobacco flavors and fragrances according to claim 1, characterized in that the number of each layer of the brackets (9) is no less than 3. 5. A method for collecting microorganisms contaminated with tobacco flavors and fragrances according to claim 1, characterized in that 30-50 mg glass beads (6) are placed in the lower half of the housing (2).