ISO 20963 pdf download – soil quality — Effects of pollutants oninsect larvae (Oxythyrea funesta)—Determination of acute toxicity

ISO 20963 pdf download – soil quality — Effects of pollutants oninsect larvae (Oxythyrea funesta)—Determination of acute toxicity.
5 Test environment
Tests 5hall be performed at a temperature of (28 ± 1) C m complete darkness.
6 Reagents
6.1 BIological material
The species used in the test is Oxythyrea fwiesta (Scarebaeidae, Cetoni.inae) Third-meter larvae with a fresh mass within the range 100 mg to 200 mg are required to perform the test. The larvae shall be healthy, without any bites or other visible injuries.
NOTE Depending on the breedwlg conditions, descxted in Annex A. larvae proxima(eIy iso weeis old are suitable for the test.
Larvae of similar size shall be selected, The difference in mass between the smallest and the largest larva within a single test container shall not exceed 50 mg.
Eliminate the particles of breeding substrate stuck to the integument using, for example, a soft brush before weighing the larvae. It Is also possible to leave the animals to move along on slightly moist paper in order to eliminate the breeding substrate stuck to the integument.
Synchronisation of breeding is necessary An example of breeding technique for Oxythyrea funesfa is given in
Annex A.
8.2 Test substrate
The mass of substrate used per glass container (7.1) shall be equivalent to 300 g (dry mass).
The substrate, called artificial soil, shall have the following composition (in accordance with ISO 11268-1):
— sphagnum peat, air-dried, finely ground and with no visible plant remnams: 10 % (expressed on a dry mass basis);
— kaol.nite clay containing not less than 30% kaolinite: 20 % (expressed on a dry mass basis):
industnal quartz sand (dominant fine sand with more than 50 % particle size between 0,05 mm and 0.20 mm): 70% (expressed on a dry mass basis).
Md pulverised calcium carbonate (CaCO3), of recognised analytical wade. as necessary to bring the p11 of the wetted substrate to 6.0±0,5 (commonly between 0.5% and I % of the mass of the dry ined.ents),
Prepare the artificial soil by mlxrng the dry constituents listed above thoroughly an a large-scale laboratory mixer The amount of calcium carbonate required can vary, depending on properties of the individual batch of sphagnum peat, and should be determined by weighing subsarnples immediately before the test.
Store the mixed artificial soil at room temperature. To determine pH and the maximum water-holding capacity. pre-moisten the dry artificial soil at least two days before starting the test by adding deionized water to obtain half of the required final water content of 50 % of the maximum water-holding capacity
8.3Preparation of control container
The control container contains the test substrate wetted with deionized water to reach 50 % of the total water-holding capacity (determined in accordance with ISO 11269-2:—, Annex A).
Prepare one control container for the preliminary test and three control containers for the final test.
lf the preparation of the test mixtures requires the use of a solvent, prepare an additional control containingthe same amount of solvent as used for introducing the test substance.Prepare the control container(s) inaccordance with 8.2.3 without addition of the test substance.
Measure the pH of the control and the control with solvent (if appropriate) in accordance with IlS0 10390.8.4Food addition
Before addition of the larvae，mix 3 g of finely ground air-dried cow-dung (6.3) with the test mixture (8.2),Proceed in the same way for control containers.
Also add food during the test in order to allow larval development. Add 2 g to 3 g of finely ground air-driedcow-dung (6.3)on the surface at day 3 and day 7 (only if the previous addition has been consumed).
8.5 Introduction of the biological material
Weigh 10 larvae individually and introduce them in each container. Cover it with a polyethylene membrane(7.5) perforated with small holes in order to allow exchange between the medium and the atmosphere and toavoid escape of larvae from the test container.
8.6Test conditions and measurements
Place the containers in the test enclosure (Clause 5) for 10 days.
After 10 days,determine for each container the total number and the mass of living larvae (individual massoptional).A larva is considered to be dead if it displays no reaction to a pinprick applied to its anterior side.Note the symptoms observed on the animals.
At the end of the test,measure the pH in one control container and in one container per test mixture inaccordance with ISO 10390.
8.7 Reference substance
In order to check the biological material used,conduct at regular intervals a test with 2,4.5-trichlorophenol(6.4) in accordance with the procedure described in 8.2.2,8.3,8.4 and 8.5.