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Diss Factsheets

Ecotoxicological information

Toxicity to soil macroorganisms except arthropods

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Administrative data

Endpoint:
toxicity to soil macroorganisms except arthropods: short-term
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
6 January 2010 - 11 February 2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report date:
2010

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 207 (Earthworm, Acute Toxicity Tests)
Deviations:
yes
Remarks:
Some of the temperatures were observed to be slightly in excess of the 20 ± 2 °C given in the study plan. However, this deviation was considered not to have affected the outcome or the validity of the test.
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Chemical structure
Reference substance name:
Calcium carbonate
EC Number:
207-439-9
EC Name:
Calcium carbonate
Cas Number:
471-34-1
Molecular formula:
CH2O3.Ca
IUPAC Name:
calcium carbonate
Test material form:
solid: nanoform, no surface treatment
Details on test material:
- Name of test material (as cited in study report): Calcium carbonate (nano)
- Description: White powder
- Batch No.: GICM014427
- Storage condition of test material: Room temperature in the dark

The nano form of calcium carbonate was tested because this form was anticipated to represent the worst case as it is likely to be more soluble than the bulk form due to the smaller particle size and hence greater surface area.
Specific details on test material used for the study:
uncoated nano CaCO3

Sampling and analysis

Analytical monitoring:
no
Details on sampling:
Not applicable

Test substrate

Vehicle:
no
Details on preparation and application of test substrate:
- Method of mixing into soil: The test item was prepared by direct addition to the basic soil substrate. An amount of test item (3500 mg) was added to approximately 100 g premix of artificial soil and carefully mixed with a spatula until homogenous. This premix was then incorporated into 3.5 kg (dry weight) of artificial soil with 1000 mL of deionised reverse osmosis water and mixed using a Hobart A200N mixer.
- Controls: The control was prepared in an identical manner using 2.5 kg (dry weight) of artificial soil and 710 mL of deionised reverse osmosis water, without the addition of test item.

Test organisms

Test organisms (species):
Eisenia fetida
Animal group:
annelids
Details on test organisms:
TEST ORGANISM
- Common name: Earthworms
- Source: Original Organics Limited, Langlands Business Park, Devon, UK
- Age at test initiation (mean and range, SD): at least 2 months old with clitellum
- Weight at test initiation (mean and range, SD): 0.44 - 0.52 g (mean 0.49 g)


ACCLIMATION
- Acclimation period: The earthworms were transferred into artificial soil (without test item) approximately 24 hours prior to the start of the test.

Study design

Study type:
laboratory study
Substrate type:
artificial soil
Limit test:
yes
Total exposure duration:
14 d
Post exposure observation period:
Earthworms were observed for up to 14 days

Test conditions

Test temperature:
19 to 23 °C
pH:
Test initiation: 5.8 - 7.1
Test termination: 6.3 - 6.6
Moisture:
The actual moisture content of the soil used in the definitive test was 29% of dry weight of basic substrate on Day 0 and 23% and 24% on Day 14.
Details on test conditions:
TEST SYSTEM
- Test container (material, size): 1 L glass beakers were used
- Amount of soil or substrate: The test vessels contained approximately 3.5 kg (dry weight) of artificial soil and the control vessels contained 2.5 kg.
- No. of organisms per container (treatment): 10 earthworms per test and control vessel
- No. of replicates per treatment group: 6
- No. of replicates per control: 4


SOURCE AND PROPERTIES OF SUBSTRATE
- Composition of the artificial soil:
Industrial quartz sand - 70% w/w
Kaolinite clay - 20% w/w
Sphagnum moss peat - 10% w/w
- Maximum water holding capacity (in % dry weight): Nominal: 30% (this level of moisture has been shown to ensure that a good crumb structure is formed within the soil).
- Stability and homogeneity of test material in the medium: No analysis was conducted to determine the homogeneity, concentration or stability of the test item formulation.


OTHER TEST CONDITIONS
- Photoperiod: Continuous illumination
- Light intensity: 684 to 714 lux


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : The weight of each earthworm was recorded on Day 0 before addition to the test or control vessels. Each surviving earthworm was weighed on Day 14.
Any mortalities were recorded 7 and 14 days after the start of the treatment and any sub-lethal effects were recorded daily. The criterion of death was taken to be the absence of any reaction to a physical stimulus at either end of the body.


TEST CONCENTRATIONS
- Range finding study
- Test concentrations: Nominal concentrations: 100 and 1000 mg/kg
- Results used to determine the conditions for the definitive study: The results showed no mortalities at the test concentrations of 100 and 1000 mg/kg.
Nominal and measured concentrations:
Definitive test:
Nominal concentration: 1000 mg/kg of soil
Reference substance (positive control):
yes
Remarks:
Chloroacetamide (nominal concentrations 5.6, 10, 18, 32 and 56 mg/kg)

Results and discussion

Effect concentrationsopen allclose all
Duration:
14 d
Dose descriptor:
LC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
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:
other: mortality and sub-lethal effects
Details on results:
- Mortality at end of exposure period: There were no significant mortalities or sub-lethal effects of exposure in 60 earthworms exposed to a test concentration of 1000 mg/kg for a period of 14 days. Two worms are unaccounted for on Day 14 in the 1000 mg/kg test concentration. This was considered to be possibly due to natural causes rather than a toxic effect given that this constituted less than 5% of the total number of earthworms exposed to this concentration and no other effects were observed (see table 1).
- Weight at end of exposure period: There were no significant differences between the control and the 1000 mg/kg test groups in terms of weight on Days 0 and 14 (see table 2).
Results with reference substance (positive control):
- Results with reference substance: The results from the positive control with chloroacetamide are within the normal range for this reference item of 10 to 50 mg/kg.
- Relevant effect levels: 7 day LC50: 44 mg/kg (95% confidence limits 42-45 mg/kg); 14 day LC50: 43 mg/kg (95% confidence limits 41-45 mg/kg); NOEC: 18 mg/kg

Any other information on results incl. tables

Table 1: Cumulative mortality data in the definitive test

Nominal concentration

(mg/kg)

Cumulative mortality (initial population: 10)

% mortality

Day 0

Day 14

Control

 -

R1

0

0

0

0

R2

0

0

0

0

R3

0

0

0

0

R4

0

0

0

0

1000

R1

0

0

0

0

R2

0

1*

0

10

R3

0

0

0

0

R4

0

0

0

0

R5

0

0

0

0

R6

0

1*

0

10

R1-R6 = Replicates 1 to 6

* Single mortality observed was considered to be possibly due to natural causes and was considered not to affect the validity of the test given that less than 5% mortality occurred at this concentration.

 

 

Table 2: Mean earthworm weights in the definitive test

Nominal concentration

(mg/kg)

Mean earthworm weight (g)

Day 0

Day 14

Mean

SD

Mean

SD

Control

 -

R1

0.51

0.09

0.51

0.09

R2

0.52

0.08

0.49

0.10

R3

0.51

0.08

0.47

0.10

R4

0.49

0.05

0.51

0.10

1000

R1

0.47

0.07

0.49

0.06

R2

0.50

0.08

0.51

0.11

R3

0.50

0.09

0.49

0.09

R4

0.51

0.07

0.53

0.09

R5

0.49

0.09

0.47

0.09

R6

0.44

0.08

0.48

0.09

R1-R6 = Replicates 1 to 6

SD = Standard deviation

 

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Remarks:
Mortality of control animals < 10%
Conclusions:
The acute toxicity of the test item, to the earthworm Eisenia foetida, in an artificial soil test gave a 14 day LC50 value of >1000 mg/kg and a NOEC of 1000 mg/kg.