3a,4,7,7a-tetrahydro-4,7-methanoindene

Toxicity to soil macroorganisms except arthropods

Link to relevant study record(s)

Reference

Endpoint:

toxicity to soil macroorganisms except arthropods: long-term

Type of information:

experimental study

Adequacy of study:

key study

Study period:

28/04/2016 - 12/12/2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 222 (Earthworm Reproduction Test (Eisenia fetida/Eisenia andrei))

Deviations:

yes

Remarks:

These deviations are considered to have no impact on quality and integrity of the study as all relevant validity criteria were met in the control group.

Principles of method if other than guideline:

Deviations
Food was provided on day 0 (test start) instead of day 1 due to good experience with this procedure.
The initial pH-value of the soil was slightly above the recommended value of 6 ± 0.5.
At the test end, the soil moisture deviated up to 34 % from the initial value in all treatments.

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Test item (as mentioned in the report): Dicyclopentadiene High Purity
Sample ID number: 1518148 (LIMS Sample manager report date 2016-02-17)
CAS RN (a.s.): 77-73-6
Active substance/Content: 92.66%
pH: Not applicable
Appearance: Colourless liquid
Water solubility <= 0.01%

Analytical monitoring:

no

Details on sampling:

At experimental starting and experimental completion, the pH value and moisture content of the test medium were determined in each treatment and the control from pooled samples of all replicates. The water content of each test container was checked weekly by weighing. The weight loss was replenished with the appropriate amount of demineralised water. The body weight of the adult earthworms was determined individually on day 0 and day 28 for each replicate.
After the first four weeks, the adult earthworms were removed.
Mortality and morphological changes of the adult earthworms were recorded for each replicate.
After a further four weeks, the number of offspring hatched from the cocoons was counted for each test container.

Vehicle:

no

Details on preparation and application of test substrate:

The maximum water holding capacity was determined and two days before the test start, the artifical soil was pre-moistened to 27% WHC max and kept covered until the start of exposure.
On day -1, the respective test item amount for each test item concentration was pipetted onto quartz sand (10 g per replicate) and mixed thoroughly. The treated portion of quartz sand was added to the premoistened artificial soil and mixed thoroughly with a household mixer. The spiked soil was stored in closed vessels under test conditions in the dark. On day 0 before introduction of the test organisms, demineralised water was added to the artificial soil to adjust the humidity to a moisture of 54 % of the maximum water holding capacity. Subsequently, the test medium was thoroughly mixed to ensure a homogenous distribution and about 600 g soil dry weight were filled into test vessels.

Test organisms (species):

Eisenia fetida

Animal group:

annelids

Details on test organisms:

Source: Organisms of the species Eisenia fetida are bred at the test facility in covered plastic vessels containing potting compost. Earthworms of homogeneous age and body weight were used in the definitive test.
Age: Adult earthworms with clitellum, aged between 2-12 months, though the difference in age of the earthworms used did not deviate by more than 1 month. Synchronisation of the population was achieved by placing adult earthworms into breeding boxes and removing the adults after 4 weeks. Offspring from the remaining cocoons reached the adult age after 2 months at the earliest.
Body weight: 0.35 to 0.59 g.
Adaptation: Earthworms were adapted to the artificial soil to be used for the test two days prior to test start.

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

no

Total exposure duration:

8 wk

Post exposure observation period:

None

Test temperature:

Nominal: 20 ± 2°C
Actual: 18 - 21.5°C

pH:

pH varied from 6.01 - 6.85 between the replicates

Moisture:

The moisture of the soil varied between 18.6 - 25.8% of dry weight

Details on test conditions:

- Test System:
Test container: 1.5 L glass dishes covered with opaque aluminium foil (foil was perforated from day 7 onwards)
No. of organisms per container: 10
No. of replicates per treatment group: 4
No. of replicates per control: 8
Feeding: 5 g air dried cattle manure was supplied weekly during the first 4 weeks. At test start and day 28, the food was mixed carefully into the soil and on other days was placed onto the soil surface.
Moistening: Test replicates were weighed weekly and demineralised water was added to maintain the soil moisture within 10% of the initial moisture value. 10 mL of water was added on day 7 and 5 mL on days 14, 21 and 28.

- Source and properties of substrate (if soil):
Soil: Artifical soil with quartz sand
% sand: 74 (air dried quartz sand with >50% particle size of 0.05-0.2 mm)
% clay: 20 kaolin (kaolinite content >30%)
% peat: 5 (air dried and finely ground)
% calcium carbonate: 0.21

- Other test conditions:
Photoperiod: 16 h light - 8 h dark
Light intensity: 400-800 Lux nominal (511 ± 86 actual)

- Effect parameters measured:
Body weights of adult earthworms were determined individually on days 0 and 28. Mortality and morphological changes of adult earthworms were recorded for each replicate on removal of the adult earthworms on day 28. The number of offspring hatched from the cocoons was counted for each container at the end of the test.

- Range finding study:
Range finding study: Yes, 1, 10, 100 and 1000 mg/kg soil dry weight

Nominal and measured concentrations:

Nominal: 15.6, 31.3, 62.5, 125, 250, 500 mg/kg soil dry weight
Control: Artificial soil with quartz sand moistened with demineralised water not containing test or reference item

Reference substance (positive control):

yes

Remarks:

Carbendazim

Key result

Duration:

56 d

Dose descriptor:

NOEC

Effect conc.:

125 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Key result

Duration:

56 d

Dose descriptor:

EC50

Effect conc.:

250 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Details on results:

- Mortality:
No evident earthworm mortality (< 10%) was observed in the control or test item concentrations 15.6 to 250 mg/kg soil dry weight (SDW). However, at the test item concentration 500 mg DCPD/kg SDW 100 % mortality was observed. The LC50 value for mortality was determined to be 329 mg/kg SDW.

- Pathology and behaviour:
No significant pathological symptoms or changes in the behaviour of adult earthworms were observed in the control or test item concentrations 15.6 to 250 mg/kg SDW. However, at the test item concentration 500 mg/kg SDW, 100% mortality was observed.

- Live weight:
At test start, the earthworms had individual weights in the recommended range of 0.35 - 0.59 g. There were no statistically significant differences in earthworm body weights in the treatment rates 15.6 to 250 mg/kg soil dry weight compared to the control.

- Reproduction:
The reproduction rate (average number of juveniles produced per replicate) was 149 in the control and ranged from 0 to 165 in the test item treatment rates. There were no statistically significant differences in earthworm reproduction in the treatment rates 15.6 to 125 mg/kg SDW compared to the control. However, at the test item concentration 250 mg/kg SDW, the earthworm reproduction was statistically significantly reduced, and in the test item concentration 500 mg/kg SDW, no juvenile were found. The EC50-value for reproduction was determined to be 250 mg/kg SDW. The coefficient of variation calculated for the reproduction of the control was 17.3% and thus lower than the 30%, required for a valid test.

Results with reference substance (positive control):

Test is conducted once every 12 months. Most recent study 21 January to 18 March 2016 (Study RBR1601). Significant effects were observed at 2 and 4 mg/kg SDW which is within the expected range of 1 - 5 mg/kg SDW.

Reported statistics and error estimates:

The arithmetic mean, NOEC and LC50 were determine for adult mortality. Significant differences in biomass and reproduction between test vessels and the control were determined by probit analysis, with Shapiro-Wilk Normality Test and Levene's Equal Variance Test (p=0.05). For reproduction, EC50, the arithmetic mean and the coefficient of variation per treatment and control were calculated.

N/A

Validity criteria fulfilled:

yes

Remarks:

Adult mortality in control = 10% in first four weeks, average number of juveniles in control was > 30 per replicate, coefficient of variation for reproduction in control was < 30% at the end of the test.

Conclusions:

The NOEC of the test item was determined to be 125 mg/kg SDW based on reproduction. The EC50 value for reproduction was determined to be 250 mg/kg SDW.

Executive summary:

The effect of DCPD on earthworm reproduction was determined in a GLP-compliant study following OECD guideline 222. Earthworms were exposed to 6 concentrations of DCPD alongside a control in artifical soil. There were no statistically significant differences in earthworm reproduction in the treatment rates 15.6 to 125 mg DCPD/kg SDW compared to the control. However, at the test item concentration 250 mg DCPD/kg, the earthworm reproduction was statistically significantly reduced and, at the test item concentration 500 mg DCPD/kg soil dry weight, no juveniles were found. Overall, the NOEC of the test item concerning mortality, biomass and reproduction was determined to be 125 mg DCPD/kg SDW and based on reproduction. The EC₅₀ value for reproduction was determined to be 250 mg DCPD/kg SDW.

Description of key information

The NOEC of the test item was determined to be 125 mg/kg SDW based on reproduction. The EC₅₀-value for reproduction was determined to be 250 mg/kg SDW. The calculated LL₅₀ from TLM QSAR towards E. fetida is 98.649 mg/kg soil dw and NOELR is 18.898 mg/kg soil dw.

Key value for chemical safety assessment

Short-term EC50 or LC50 for soil macroorganisms:

250 mg/kg soil dw

Long-term EC10, LC10 or NOEC for soil macroorganisms:

125 mg/kg soil dw

Additional information

The effect of DCPD on earthworm reproduction was determined in a GLP-compliant study following OECD guideline 222. Earthworms were exposed to 6 concentrations of DCPD alongside a control in artificial soil. There were no statistically significant differences in earthworm reproduction in the treatment rates 15.6 to 125 mg DCPD/kg SDW compared to the control. However, at the test item concentration 250 mg DCPD/kg, the earthworm reproduction was statistically significantly reduced and, at the test item concentration 500 mg DCPD/kg soil dry weight, no juveniles were found. Overall, the NOEC of the test item concerning mortality, biomass and reproduction was determined to be 125 mg DCPD/kg SDW and based on reproduction. The EC₅₀ value for reproduction was determined to be 250 mg DCPD/kg SDW.