2H-Pyran, 2-[4'-[difluoro(3,4,5-trifluorophenoxy)methyl]-3',5'-difluoro[1,1'-biphenyl]-4-yl]-5-ethyltetrahydro-

Toxicity to soil macroorganisms except arthropods

Link to relevant study record(s)

Reference

Endpoint:

toxicity to soil macroorganisms except arthropods: short-term

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2009-03-06 till 2009-03-20

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 207 (Earthworm, Acute Toxicity Tests)

Version / remarks:

adopted April 4, 1984

Deviations:

no

Qualifier:

according to guideline

Guideline:

ISO 11268-1 (Effects of Pollutants on Earthworms. 1. Determination of Acute Toxicity Using Artificial Soil Substrate)

Version / remarks:

1997

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

yes

Remarks:

Acetone

Details on preparation and application of test substrate:

The test item was dissolved in acetone and mixed with quartz sand. After evaporation of the acetone the treated quartz sand was incorporated into the artificial soil.

Test organisms (species):

Eisenia fetida

Animal group:

annelids

Details on test organisms:

TEST ORGANISM
- Common name: Earthworm
- Source: Breeding at DR. U.NOACK-LABORATORIEN
- Age at test initiation: adult (with cliteilum)
- Weight at test initiation: 0.3 to 0.6 g
- Breeding: Organisms of the species Eisenia fetida are bred at the test facility in covered plastic vessels (volume 60 L) containing potting compost at 20°C in the dark.
- Feeding: The earthworms are fed with dried litter of stinging nettle and porridge oats. The amount of the food is portioned due to the feeding rate and due to the density of worms in the vessels.

ACCLIMATION
- Acclimation period: yes, for two days
- Acclimation conditions: same as test conditions
- Health during acclimation: no mortality observed

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

yes

Total exposure duration:

14 d

Test temperature:

18.0-22.0 °C

pH:

4.98 - 5.28

Moisture:

32.7 - 34.4 % of dry weight

Details on test conditions:

TEST SYSTEM
- Test container: Glass dishes (volume 1.5 L), closed with perforated plastic film
- Amount of soil or substrate:
- No. of organisms per container (treatment): 10
- No. of replicates per treatment group: 4
- No. of replicates per control: 4
- No. of replicates per vehicle control: 4

SOURCE AND PROPERTIES OF SUBSTRATE
Artificial soil consisted of the following components:
-10 % peat (air dried and finely ground)
- 20 % kaolin (kaolinite content 84 - 87 %)
- 69 % quartzsand (sand with > 50 % particles size of 0.05 to 0.2 mm)
- 0.35 % calcium carbonate (CaC03) to adjust the pH to 6.0 ± 0.5
- Maximum water holding capacity: 66 g /100 g dry weight

OTHER TEST CONDITIONS
- Photoperiod: 24h
- Light intensity: 568 ± 85.8 lux

EFFECT PARAMETERS MEASURED
The live weight of the earthworms was measured individually at the day of application (day 0) and at day 14. Mortality, behaviour and morphological changes of the earthworms were determined 7 and 14 days after application. At start and end of the test, pH-value and moisture content of the test medium were determined in the treatment and controls using mixed samples of all four replicates.

VEHICLE CONTROL PERFORMED: yes

TEST CONCENTRATIONS
- Range finding study: yes
- Test concentrations: 10, 100, 1000 mg test item/kg soil dry weight
- Results used to determine the conditions for the definitive study: yes, no mortality occured and the no biomass decrease was induced

Nominal and measured concentrations:

Nominal concentration: 1000 mg/kg soil dry weight
no measured concentration

Reference substance (positive control):

yes

Remarks:

2-Chloroacetamide

Key result

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Details on results:

- Mortality at end of exposure period: One dead earthworm was found in the control after seven days of exposure, which is considered to be an artefact. In the solvent control and at the limit test item concentration of 1000 mg/kg soil dry weight no earthworm mortality occurred after 14 days of exposure.
- Changes in body weight of live adults (% of initial weight) at end of exposure period: Control: 6% loss, solvent control: 4% loss, test item: 3 % loss
- Morphological abnormalities: no
- Behavioural abnormalities: no
- Other biological observations: no
Validation criteria:
The mortality in the control groups should not exceed 10 % at the end of the test. The average loss of biomass of the worms in the controls should not exceed 20 %. Both criteria are fulfilled.

Results with reference substance (positive control):

The acute toxicity of 2-Chloroacetamide on the earthworm Eisenia fetida fetida (SAVIGNY) was determined over a period of two weeks from February 12th to 26th, 2009. The LC50-value ranged between 20 and 80 mg/kg soil dry weight as recommended by the guideline. LD50: 50.1 mg/kg soil dry weight (95% - Confidence Interval: 45.8 - 55.5); NOEC 30.0 mg/kg soil dry weight

Reported statistics and error estimates:

Differences between the earthworm biomass in the control and the solvent control were statistically analysed using a t- test. Due to no statistical differences both controls were pooled for further statistical analyses.t-tests were carried out for the determination of statistically significant differences compared to the pooled control. The α-value (acceptable probability of incorrectly concluding that there is no difference) was α=0.05. Calculations were carried out using software
- Excel, MICROSOFT
- Sigma Stat, SPSS INC.
- Sigma Plot, SPSS CORPORATION

Table 1: Mean biomass and mean loss of body weight

Nominal test item concentration [mg/kg soil dry weight]	Mean live weight			Comparison versus Control
	Test start MV±SD [g/worm]	Test end MV±SD [g/worm]	Test end cv[%]	Loss [%I	Significance
Contro	0509±0.062	0.480±0.075	15.5	6	-
Solvent Control	0 511±0.060	0.490±0.056	11.4	4
1000	0.510±0.064	0.494±0.069	14.0	3	No

MV: mean value

CV: coefficient of variation

SD: standard deviation

Validity criteria fulfilled:

yes

Conclusions:

In the test according to OECD 207 the toxicity to earthworm in artificial soil was higher than 1000 mg/kg soil dry weight, the NOEC is 1000mg/kg soil dry weight.

Executive summary:

The acute effects of the test item on the earthworm Eisenia fetida were determined according to the OECD Guideline 207 and DIN ISO 11268-1. The study was conducted with a limit test item concentration of 1000 mg/kg soil dry weight throughout 14 days. Artificial soil without test item was used as control and artificial soil without test item but treated with the solvent was used as solvent control. 40 test organisms with an individual weight between 0.3 -0.6 g, divided into four replicates, were tested at the limit test item concentration and controls in artificial soil containing 10 % peat. No mortality as well as no pathological symptoms or changes in the earthworm behaviour were observed at the limit test item concentration of 1000 mg/kg soil dry weight after 14 days of exposure.One dead earthworm was found in the control after seven days of exposure, which is considered to be an artefact.Compared to the pooled control, the biomass of the earthworms at the limit test item concentration of 1000 mg/kg soil dry weight was not statistically significantly reduced at test end. Consequently, the no observed effect concentration (NOEC) with respect to mortality, loss of biomass and other symptoms was determined to be at 1000 mg/kg soil dry weight. All validity criteria of the test guideline were fulfilled.

Description of key information

In the test according to OECD 207 the toxicity to earthworm in artificial soil was higher than 1000 mg/kg soil dry weight, the NOEC is 1000mg/kg soil dry weight (reference 6.3.1 -1).

Additional information

The acute effects of the test item on the earthworm Eisenia fetida were determined according to the OECD Guideline 207 and DIN ISO 11268-1. The study was conducted with a limit test item concentration of 1000 mg/kg soil dry weight throughout 14 days. Artificial soil without test item was used as control and artificial soil without test item but treated with the solvent was used as solvent control. 40 test organisms with an individual weight between 0.3 -0.6 g, divided into four replicates, were tested at the limit test item concentration and controls in artificial soil containing 10 % peat. No mortality as well as no pathological symptoms or changes in the earthworm behaviour were observed at the limit test item concentration of 1000 mg/kg soil dry weight after 14 days of exposure.One dead earthworm was found in the control after seven days of exposure, which is considered to be an artefact.Compared to the pooled control, the biomass of the earthworms at the limit test item concentration of 1000 mg/kg soil dry weight was not statistically significantly reduced at test end. Consequently, the no observed effect concentration (NOEC) with respect to mortality, loss of biomass and other symptoms was determined to be at 1000 mg/kg soil dry weight. All validity criteria of the test guideline were fulfilled (reference 6.3.1 -1).