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Method for detecting influence of buccal tobacco product on micronucleus rates of cells

A technology for detecting snus products and detection ports, which is applied in the field of detecting the influence of snus products on the micronucleus rate of cells, and can solve problems such as the inability to meet the needs of genotoxicity detection of snus products

Active Publication Date: 2015-07-01
CHINA TOBACCO YUNNAN IND
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Problems solved by technology

The use of snus products is different from that of traditional cigarette inhalation. The snus products are used directly by mouth, and their dissolved substances will enter the digestive system with saliva. During use, no smoke is produced, and the exposure route is significantly different from traditional cigarettes. For cigarettes, changes in the test object lead to different sample pretreatment methods, test cells, and test doses. The in vitro micronucleus test used to test the genotoxicity of traditional cigarettes cannot meet the needs of genotoxicity testing for snus products. Research institutes and tobacco companies have not formulated a standard method for in vitro micronucleus testing of snus products. Therefore, it is necessary to establish an in vitro micronucleus test method suitable for snus products according to their own characteristics to ensure product safety.

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  • Method for detecting influence of buccal tobacco product on micronucleus rates of cells

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Embodiment 1

[0037] It is used for the detection of genotoxicity of Camel brand mint flavored Snus snus products.

[0038](1) Sample pretreatment method: Accurately weigh 1.0g of Camel brand mint-flavored Snus snus product, add 30mL of commercially available serum-free medium, and place it in a constant temperature water bath at 37°C for 24h for extraction. After extraction, use qualitative filter paper for primary filtration to remove slag, and then use 0.45 μm organic filter membrane to filter and sterilize to obtain the test solution;

[0039] (2) Preparation of single cell suspension: hOMF cells of human oral mucosal fibroblasts were incubated at 37°C, 5% CO 2 , 95%RH (relative air humidity) incubator, observe the confluence and morphology of the cultured cells through an inverted microscope, remove the medium when the confluence of the cells is about 80-90%, wash twice with phosphate buffer, discard Wash solution; add 1mL-2mL trypsin solution with a concentration of 0.25% (w / v) for m...

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Abstract

The invention provides a method for detecting influence of a buccal tobacco product on micronucleus rates of cells. The method comprises the following steps of performing a sample pretreatment method, performing preparation of single-cell suspension, calculating cell concentration, performing cell inoculation, adding a test object, incubating the test object, harvesting the cells, performing hypotension, dropping to a slider, dyeing, performing micronucleus calculation, performing result determination and the like. According to the method provided by the invention, by comprehensively surveying the exposure way and the action mode of the buccal tobacco product, the sample pretreatment method for the buccal tobacco product is established; moreover, human oral mucosal fibroblasts hOMF are selected according to action target cells of the buccal tobacco product to serve as cells for in vitro micronucleus test detection on the buccal tobacco product, and the sample detection dosage is determined, and therefore an in vitro micronucleus test detection method suitable for the buccal tobacco product is formed.

Description

technical field [0001] The invention relates to a method for detecting the influence of snus products on the micronucleus rate of cells, and belongs to the technical field of biological safety evaluation of tobacco and tobacco products. Background technique [0002] According to the different genetic endpoints detected, more than 200 short-term detection methods of genotoxicity have been established [1]. Cellular micronucleus rate is one of the key combination indicators for genotoxicity evaluation. Due to its clear detection endpoint, simple method, and easy to carry out, micronucleus test is used in the safety evaluation of food, pharmaceutical products, industrial and agricultural products, and other health-related products. , monitoring of genetic damage, mutation detection and early prediction of specific populations have been widely used [2-5]. In 2009, the Organization for Economic Co-operation and Development (OECD) published a guidance document for in vitro micronu...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/50
CPCG01N33/5014
Inventor 李雪梅管莹夭建华高茜陈建华杨叶昆米其利朱洲海
Owner CHINA TOBACCO YUNNAN IND
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