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Biological measuring method for dioxins compound in monitoring environment samples

A dioxin-like, environment-monitoring technology, which is applied in the field of dioxin-like pollutants, can solve problems such as difficult operation of the heart infusion method, contamination of the target tissue, time-consuming and labor-intensive problems, and achieves simple operation, small sample volume, and shortened operation time Effect

Inactive Publication Date: 2004-11-24
RES CENT FOR ECO ENVIRONMENTAL SCI THE CHINESE ACAD OF SCI
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Problems solved by technology

[0005] The purpose of the present invention is to overcome the difficult operation of the heart infusion method used in the prior art to test dioxin-like pollutants using primary cultured hepatocyte technology, which is time-consuming and labor-intensive, and is likely to cause contamination of the target tissue, and the fish species used It is not applicable to fish species in my country, so as to provide a cost-effective biomonitoring method for dioxin-like compounds. Grass carp, the most common fish species in my country, is used to detect dioxin-like compounds in environmental samples by using its primary cultured liver cells. Dioxin-like pollutants, and the content of dioxin-like pollutants in environmental samples can be calculated through the standard curve

Method used

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  • Biological measuring method for dioxins compound in monitoring environment samples
  • Biological measuring method for dioxins compound in monitoring environment samples
  • Biological measuring method for dioxins compound in monitoring environment samples

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Example 1: Using the primary culture method of grass carp hepatocytes to measure the toxic effect of TCDD to draw a standard curve

[0020] Primary cell culture: Disinfect 1.0Kg of grass carp to be tested, remove the blood from the tail vein, dissect the fish body to take a small piece of liver tissue, and use 0.145mol / L NaCl, 5.4mmol / L KCl, 12mmol / L NaHCO 3 , 5mmol / L Na 2 EDTA, 3mmol / L NaH 2 PO 4 , 1.1mmol / L KH 2 PO 4 , 100mmol / L N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid (hereinafter referred to as HEPES) buffer solution (pH7.5) washed to white. Cut the liver tissue into 1mm 3After size, digest with 0.25v% trypsin digestion solution 30 times the weight of the liver for 2 to 3 minutes, and stop the digestion when the cells of the liver tissue are dispersed into small cell clusters; pass through a 100-mesh sieve, transfer the filtrate into a centrifuge tube, Centrifuge at 1000r / min for 2min, pour off the supernatant, and wash the pellet with DMEM / F12 mediu...

Embodiment 2

[0024] Example 2: Using the primary culture method of grass carp liver cells to evaluate the toxicity of dioxin-like pollutants in environmental samples

[0025] River sediment, electrolytic waste residue, floating dust, sediment standard sample, standard soil and chemical products were collected as samples. Using Soxhlet extraction method, adding toluene to extract dioxin-like pollutants in the sample, loading the extracted organic solvent on a silica gel / activated silica gel column to purify the sample, and then eluting the concentrated component with n-hexane , Blow dry in high-purity nitrogen, add DMSO to dissolve and prepare a test sample solution with any concentration within the linear range of the standard curve.

[0026] Utilize the primary culture method of grass carp liver cells described in Example 1 to evaluate the toxicity of dioxin-like pollutants in environmental samples, and the measurement results are compared with those of the H4IIE cell culture method and h...

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Abstract

The present invention is the method of monitoring dioxins pollutant in environment sample with grass carp liver cell obtained via primary culture technology. Grass carp is blood eliminated via tail vein and its liver cell is primary cultured; the liver cell is then cultured together with 2, 3, 7, 8-tetrachlorodibenzyl dioxins; reactant solution containing excessive 7-oxethyl-isophenol azolactone is then added; the fluorescent strength of the end product 7-hydroxy-isophenol azolactone is measured with fluorophotometer and the fluorescent strength is used as parameter for expressing EROD enzyme activity to draw the standard curve of dosage of EROD enzyme activity of primary liver cell vs 2, 3, 7, 8-tetrachlorodibenzyl dioxins. The environment sample is detected similarly and through the comparison with standard curve the dioxins pollutant content in environment sample may be calculated.

Description

technical field [0001] The invention relates to a biological testing method, in particular to a method for monitoring dioxin-like pollutants in environmental samples by using grass carp liver cells obtained by primary culture technology. technical background [0002] Dioxin-like compounds include dioxins, furans, polychlorinated biphenyls, and other coplanar halogenated aromatics, which are characterized by aryl hydrocarbon receptor effects. These compounds are highly carcinogenic, teratogenic, mutagenic, interfere with nervous / endocrine system effects, and persist in the environment. They have become key pollutants of global concern in the environmental field. To monitor dioxin-like pollutants in environmental samples, chemical analysis and biological testing techniques are usually used. Chemical analysis requires the use of high-resolution chromatography / high-resolution mass spectrometry, or chromatography / isotope dilution mass spectrometry, so the analysis cost is relati...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/02G01N21/64
Inventor 王子健万小琼马陶武吴文忠
Owner RES CENT FOR ECO ENVIRONMENTAL SCI THE CHINESE ACAD OF SCI
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