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GB/T 31270 consists of 21 parts under the general title Test guidelines on environmental safety assessment for chemical pesticides:
——Part 1: Transformation in soils;
——Part 2: Hydrolysis;
——Part 3: Phototransformation;
——Part 4: Adsorption/desorption in soils
——Part 5: Leaching in soil;
——Part 6: Volatility;
——Part 7: Bioconcentration test;
——Part 8: Degradation in water-sediment systems;
——Part 9: Avian acute toxicity test;
——Part 10: Honeybee acute toxicity test;
——Part 11: Silkworm acute toxicity test;
——Part 12: Fish acute toxicity test;
——Part 13: Daphnia sp.acute immobilisation test;
——Part 14: Alga growth inhibition test;
——Part 15: Earthworm acute toxicity test;
——Part 16: Soil microorganism toxicity test;
——Part 17: Trichogramma acute toxicity test;
——Part 18: Amphibian acute toxicity test;
——Part 19: Effects on non-target plants;
——Part 20: Livestock short-term dietary toxicity test;
——Part 21: Macro-crustacean toxicity test.
This is Part 15 of GB/T 31270.
This part is developed in accordance with the rules given in GB/T 1.1-2009.
Attention is drawn to the possibility that some of the elements of this document may be the subject of patent rights. The issuing body of this document shall not be responsible for identifying any or all such patent rights.
This standard was proposed by and is under the jurisdiction of the Ministry of Agriculture of the People's Republic of China.
Test guidelines on environmental safety assessment for chemical pesticides - Part 15: Earthworm acute toxicity test
1 Scope
This part of GB/T 31270 specifies the basic requirements for materials, conditions, operation, quality control, data processing, test reports, etc. for acute toxicity test of chemical pesticides on earthworm.
This part is applicable to earthworm acute toxicity test carried out for the registration of chemical pesticides. It may be used as reference for other types of pesticides.
This part is not applicable to volatile and insoluble chemical pesticides.
2 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
2.1
clitellum
gland on the epidermis of the front of the earthworm, which is saddle-shaped or band-shaped and is usually separated from the rest of the body by color
2.2
adult worm
earthworm with clitellum in the front of the body
2.3
median lethal concentration
concentration of the test substance at which 50% of the test organisms are killed in the acute toxicity test, expressed by LC50
Note: The unit is in mg a.i./kg dry soil.
2.4
test substance
substance to be tested
2.5
chemical pesticide
pesticide made from chemical substances through artificial synthesis, some of which are synthetic bionic pesticides made through imitation, structural transformation and innovation by taking the active substances in natural products as the matrix
Synonym: synthetic organic pesticide.
[Definition 2.3.1, NY/T 1667.1-2008]
2.6
technical material
final product consisting of the active ingredients and impurities obtained during the manufacturing process, which shall not contain visible foreign substances or any additives, and if necessary, a small amount of stabilizer may be added
[Definition 2.5.1, NY/T 1667.2-2008]
2.7
formulation product
product stable in use and made from technical materials (or technical concentrate) of pesticides and auxiliaries
[Definition 2.4, NY/T 1667.2-2008]
2.8
active ingredient; a.i.
biologically active ingredient (with a specific chemical structure) in pesticide products
[Definition 3.1, NY/T 1667.2-2008]
2.9
reference substances
chemical substance or mixture used in a test to confirm or deny certain characteristics of a test substance or to judge the effectiveness of a test system
3 Test overview
Add the pesticide solution to an appropriate amount of artificial soil and mix well, and place 10 earthworms in each treatment group and cultivate them for two weeks under suitable conditions. Observe and record the toxicity symptom and number of dead earthworms at the 7d and the 14d, and then work out the median lethal concentration LC50 and 95% confidence limits of pesticides to earthworms.
4 Test methods
4.1 Materials and conditions
4.1.1 Test organism
It is recommended to use the adult worm of Eisenia foetida which are 0.30 g and 0.60 g in weight for the test.
4.1.2 Test soil
Artificial soil (see Annex A for formulations).
4.1.3 Test substance
Formulation products, technical materials or pure pesticides shall be used as test substance. Insoluble test substances can be dissolved with a small amount of organic solvent which is less toxic to earthworms, and in general, the amount thereof shall not exceed 0.1mL (g)/L.
4.1.4 Main apparatus and instruments
Main apparatus and instruments are as follows:
- incubator;
- specimen bottle;
- volumetric flask, etc.
4.1.5 Test conditions
The test shall be carried out at a temperature of 20±2℃, a relative humidity of 70%~90%, and a illumination intensity of 400lx to 800lx.
4.2 Test operations
4.2.1 Pre-test
Set several groups of concentrations at a large difference according to the formal test conditions so as to calculate the lowest concentration of the test substance at which all earthworms are killed and the highest concentration the test substance at which all earthworms are survived, and hereby set the concentration for formal test.
4.2.2 Formal test
Set 5~7 concentration groups at a certain differential within the concentration range determined by the pre-test, and set a blank control group. If cosolvent is used, a solvent control group shall also be added, and a blank control without pesticide shall be arranged, with 3 replicates set for each concentration group. Place 500g of soil in the specimen bottle (with the soil thickness in the specimen bottle no less than 8cm), add pesticide solution and mix well (if the organic solvent is used to dissolve the solvent, the organic solvent shall be fully volatilized), and add an appropriate amount of distilled water to adjust the moisture content of the soil, so that it accounts for 30% to 35% of the dry weight of the soil. Put 10 earthworms into each treatment group, tie up the bottleneck with gauze, and place the specimen bottle in an incubator with a temperature of 20℃±2℃, a humidity of 70% to 90%, and a illumination intensity of 400 lx to 800 lx. The test lasts for two weeks. Pour out the soil in the bottle at the 7d and 14d, then observe and record the toxicity symptom and the death count of earthworms (touch the end of earthworm gently with a needle, if the earthworm does not response, it is regarded as dead), and the dead earthworm shall be removed in time. And then work out LC50 and 95% confidence limits of toxicity of pesticide to earthworm according to the death rate of earthworms at 7d and 14d.
Foreword i
1 Scope
2 Terms and definitions
3 Test overview
4 Test methods
5 Test report
Annex A (Informative) Composition and ratio of artificial soil
Annex B (Informative) Classification of toxicity levels of pesticides to earthworm
Bibliography
