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Short-term toxicity to aquatic invertebrates

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Reference
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From 2000-02-04 to 2000-05-29
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
This study was performed according to international guideline with GLP statement. All validity criteria were fulfilled and no deviation from guidelines were observed. However, based on the analytical method used, this study is considered reliable with restrictions.
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Deviations:
no
Principles of method if other than guideline:
Not applicable.
GLP compliance:
yes (incl. certificate)
Remarks:
1999-09-22
Analytical monitoring:
yes
Details on sampling:
- Concentrations: 0, 0.313, 0.625, 1.25, 2.5, 5, and 10 mg/L
- Sampling method: 20 daphnids (in 4 groups of 5 each) were exposed to each concentration for 48 hours and immobilization was recorded at 0, 24, and 48 hours.
- Sample storage conditions before analysis: none
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Test solution were prepared by further dilution of the stock solution with water to provide a geometric series of concentrations. Test vessels containing daphnids were filled directly from the test solution containers immediately after preparation. Solutions were changed at T24 hours.
- Eluate: M4 reconstituted water
- Differential loading: no data
- Controls: M4 without test substance
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: daphnid
- Strain: STRAUS-clone 5
- Source: Centre technique du bois et de l'ameublement (Paris, France)
- Age at study initiation (mean and range, SD): 6-24 hours
- Weight at study initiation (mean and range, SD): no data
- Length at study initiation (length definition, mean, range and SD): no data
- Valve height at study initiation, for shell deposition study (mean and range, SD): no data
- Peripheral shell growth removed prior to test initiation: no data
- Method of breeding: no data
- Feeding during test: no

ACCLIMATION
- Acclimation period: none
Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Remarks on exposure duration:
none
Post exposure observation period:
no data
Hardness:
238-239 mg/L as CaCO3
Test temperature:
18.6-19.8°C
pH:
7.73-8.42
Dissolved oxygen:
6.3-9.1 mg O2/L
Salinity:
not applicable.
Nominal and measured concentrations:
In the definitive test:
- nominal conc.: 0, 0.313, 0.625, 1.25, 2.5, 5, and 10 mg/L
- measured conc. (5 and 10 mg/L nominal): 1.64, 2.02 mg/L
See table 6.1.3/2 in "Any other information on results incl. tables".
Details on test conditions:
TEST SYSTEM
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: 50 mL
- Aeration: no
- Renewal rate of test solution (frequency/flow rate): each 24 h
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: M4 reconstituted water

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16:8
- Light intensity: no data

TEST CONCENTRATIONS
- Range finding study: yes
- Test concentrations: a limit test at 10 mg/L (limit of solubility) and a range-finding test at 0.01, 0.1 and 1 mg/L were performed.
- Results used to determine the conditions for the definitive study: After 48 hours in the limit test, immobilizations were 0% and 15% in the control and at 3.16 mg/L (10 mg/L nominal), respectively. At the end of the test in the range-finding test, immobilizations were 0%, 0% and 5% at the nominal concentrations of 0.01,0.1 and 1 mglL, respectively.
Reference substance (positive control):
no
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 3.16 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
other: The highest concentration without observed effect
Effect conc.:
2.02 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
See tables in "Any other information on results incl. tables".

The slight difference of toxicity at 10 mg/L (nominal) between the preliminary (15% immobilization) and definitive tests (0% immobilization) could be explained by a higher level of test substance in the limit test solution than in the definitive test solution (based on geometric means of measured concentrations in these solutions, 3.16 mg/L versus 2.02 mg/L).
In this study, according to the results of the preliminary and definitive tests: the 48-hour EC50 of the test substance and the lowest concentration resulting in 100% immobilization at 48 hours were > 3.16 mg/L; the highest concentration resulting in no immobilization at 48 hours was 2.02 mg/L.
Results with reference substance (positive control):
not applicable
Reported statistics and error estimates:
no data

Table 6.1.3/1: Daphnids' immobilization at each nominal concentration in the definitive test (mg/L).

Nominal concentration

(mg/L)

Immobilization

0 h

24 h

48 h

0

0

0

0

0.313

0

0

0

0.625

0

0

0

1.25

0

0

1

2.5

0

0

0

5

0

0

0

10

0

0

0

Table 6.1.3/2: Analytical measurements

Nominal (mg/L)

Measured (mg/L)

Geometric mean (mg/L)

0h

23.9h

24h

48h

Preliminary test

10

4.52

2.45

3.56

2.25

3.16

Definitive test

5

10

2.10

3.41

1.21

1.01

2.19

4.70

1.29

1.03

1.64

2.02

Validity criteria fulfilled:
yes
Conclusions:
No immobilization was observed at any of the definitive concentration tested. According to the results of the preliminary and definitive tests: the 48-hour EC50 of the test substance and the lowest concentration resulting in 100% immobilization at 48 hours were > 3.16 mg/L, and the highest concentration resulting in no immobilization at 48 hours was 2.02 mg/L.
Executive summary:

The experiment was conducted in order to asses the effect of the test substance on Daphnia magna, following international guidelines, and under GLP.

A preliminary test which included a limit (at 10 mg/L) and a range-finding test (at 0.01, 0.1 and 1 mg/L) preceded the definitive test. The results show that in the limit test, mortality was 0% in the control and 15% in the limit test solution (10 mg/L, nominal) at the end of the 48 hour; and in the range-finding test, immobilizations were 0%, 0% and 5% at the nominal concentrations of 0.01, 0.1 and 1 mglL, respectively.

In the definitive test, six different concentrations of test substance were used together with a control at 0, 0.313, 0.625, 1.25, 2.5, 5, and 10 mg/L. 20 daphnids (in four groups of five each) were exposed to each concentration for 48 hours and immobilization was recorded at 0, 24, 48 hours. Test solutions were changed each 24 hours.

Chemical analysis revealed that measured concentrations at nominal concentration of 10 mg/L were in the range 3.41 to 4.70 mg/L at T0 and T24 hours. Similarly, for the nominal concentration of 5 mg/L, the results were in the range of 1.21 to 2.19 mg/L. According, all effects are based on geometric means of measured concentrations throughout the test, calculated as 3.16 mg/L for the limit test solution and as 1.64 and 2.02 mg/L for the definitive test solutions at 5 and 10 mg/L, respectively.

No immobilization was observed at any of the definitive concentration tested. The slight difference of toxicity at 10 mg/L (nominal) between the preliminary (15% immobilization) and definitive tests (0% immobilization) could be explained by a higher level of test substance in the limit test solution than in the definitive test solution (based on geometric means of measured concentrations in these solutions, 3.16 mg/L versus 2.02 mg/L).

In this study, according to the results of the preliminary and definitive tests: the 48-hour EC50 of the test substance and the lowest concentration resulting in 100% immobilization at 48 hours were > 3.16 mg/L, and the highest concentration resulting in no immobilization at 48 hours was 2.02 mg/L.

Description of key information

OECD Guideline 202, EU Method C.2, GLP, key study, validity 2:

48h-EC50 (Daphnia magna) > 3.16 mg/L based on geometric mean measured concentrations.

Key value for chemical safety assessment

Additional information

One key study, performed by CIT laboratory, according to international guidelines with GLP standards is available on the aquatic invertebrates (Daphnia magna).

A preliminary test which included a limit (at 10 mg/L) and a range-finding test (at 0.01, 0.1 and 1 mg/L) preceded the definitive test. The results show that in the limit test, mortality was 0% in the control and 15% in the limit test solution (10 mg/L, nominal) at the end of the 48 hour; and in the range-finding test, immobilizations were 0%, 0% and 5% at the nominal concentrations of 0.01, 0.1 and 1 mg/L, respectively.

In the definitive test, six different concentrations of test substance were used together with a control at 0, 0.313, 0.625, 1.25, 2.5, 5, and 10 mg/L. 20 daphnids (in four groups of five each) were exposed to each concentration for 48 hours and immobilization was recorded at 0, 24, 48 hours. Test solutions were changed each 24 hours.

All effects are based on geometric means of measured concentrations throughout the test, calculated as 3.16 mg/L for the limit test solution and as 1.64 and 2.02 mg/L for the definitive test solutions at 5 and 10 mg/L, respectively.

No immobilization was observed at any of the definitive concentrations tested. The slight difference of toxicity at 10 mg/L (nominal) between the preliminary (15% immobilization) and definitive tests (0% immobilization) could be explained by a higher level of test substance in the limit test solution than in the definitive test solution (based on geometric means of measured concentrations in these solutions, 3.16 mg/L versus 2.02 mg/L).

In this study, according to the results of the preliminary and definitive tests: the 48-hour EC50 of the test substance at 48 hours was > 3.16 mg/L, and the highest concentration resulting in no immobilization at 48 hours was 2.02 mg/L.