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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 January 30, 2018 to February 01, 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
not specified
GLP compliance:
yes (incl. QA statement)
Analytical monitoring:
no
Details on sampling:
No confirmatory analysis of the test solutions was performed as a validated analytical method could not be developed.
Vehicle:
no
Details on test solutions:
Preparation of test solutions
The study was run with a dilution water control and nominal exposure concentrations of 6.25, 12.5, 25, 50 and 100 mg/L. A primary stock concentrate of test substance, with a nominal concentration of 100 mg/L, was prepared by weighing a nominal 0.2 g (actual weight: 0.20001 g) of test substance into a 2000 mL volumetric flask and made up to volume with culture medium. The stock was then stirred for 13 minutes. The resultant stock was observed to be clear and colourless with no particles visible and was used to prepare the test solutions. This was achieved by the direct addition of the appropriate amount of concentrate to dilution water in a volumetric flask. The control consisted of culture medium only. In all cases the final solutions contained nutrients. The 100 mg/L stock was used directly to fill the nominal 100 mg/L concentration test vessels. All test solutions were observed as clear and colourless after approximately 15 minutes stirring. The appropriate test solution (100 mL volume) was dispensed to each test and blank vessel.
Test organisms (species):
Daphnia magna
Details on test organisms:
Test organism
The test organism was the freshwater crustacean, Daphnia magna, obtained from continuous laboratory cultures held at Scymaris. The stock cultures of D. magna were maintained in a reconstituted water medium, the same as the test dilution water, at a temperature of 20 ± 2°C. The cultures were maintained in 2 L glass vessels with a working volume of 1.6 L. A photoperiod of 16 h light: 8 h dark, with 20-minute transition periods was provided. The D. magna cultures were fed on a mixed algae diet of Chlorella vulgaris, strain CCAP 211/12 and Pseudokirchneriella subcapitata, strain CCAP 278/4. The D. magna cultures were fed daily ad libitum depending on age and density of the culture. Culture conditions were such that the D. magna reproduction was by diploid parthenogenesis. D. magna <24 h old, obtained from a single culture vessel, were used for testing. The parent animals were 11 ± 1 d old and had been maintained with a twice weekly renewal of reconstituted water medium since birth. The test organisms and the cultures from which they were obtained showed no evidence of disease before the test period. The reconstituted water medium used for testing (and maintenance of stock cultures) was Elendt's M4 D. magna medium.
Test type:
static
Water media type:
other: Elendt's M4 D. magna medium
Limit test:
no
Total exposure duration:
48 h
Test temperature:
20 ± 1ºC
pH:
7.85 to 8.27
Dissolved oxygen:
8.93 to 9.22 mg/L
Nominal and measured concentrations:
0, 6.25, 12.5, 25, 50 and 100 mg/L (nominal)
No confirmatory analysis of the test solutions was performed as a validated analytical method could not be developed.
Details on test conditions:
Test apparatus
Glass beakers of 250 mL nominal capacity were used as test vessels, with four replicates per test concentration. Each vessel contained 200 mL of test solution providing a depth of approximately 60 mm. The beakers were covered with loose fitting glass lids. The positions of the treatments were randomly allocated within the test area.
Reference substance (positive control):
not specified
Key result
Duration:
24 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Key result
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
ca. 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Key result
Duration:
48 h
Dose descriptor:
LOEC
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
There was no immobility observed in the dilution water control or the test concentrations.

Results : Biological data

Daphnia magna response

Time

(h)

Nominal concentration of test substance

 

(mg/L)

Number immobilised per replicate

Total number tested

Total number immobilised

Percentage immobilised

A

B

C

D

24

Control

0

0

0

0

20

0

0

6.25

0

0

0

0

20

0

0

12.5

0

0

0

0

20

0

0

25

0

0

0

0

20

0

0

50

0

0

0

0

20

0

0

100

0

0

0

0

20

0

0

48

Control

0

0

0

0

20

0

0

6.25

0

0

0

0

20

0

0

12.5

0

0

0

0

20

0

0

25

0

0

0

0

20

0

0

50

0

0

0

0

20

0

0

100

0

0

0

0

20

0

0


The results obtained (based on mean measured concentrations of test substance) were:

Time

EC50

95% confidence limits

Calculation method

24 h

>100 mg/L

N/A

Direct observation of the data

48 h

>100 mg/L

N/A

Direct observation of the data

Based on immobility compared to the control, the 48 h No Observed Effect Concentration (NOEC) was determined to be 100 mg/L and the Lowest Observed Effect Concentration (LOEC) was >100 mg/L. There was no immobility observed in the dilution water control or the test concentrations.

Validity criteria

The OECD 202 Test Guideline details the following performance criteria for the test validity:

1)    In the control, no more than 10% of the daphnids should have been immobilised or show other signs of stress;

2)    The dissolved oxygen concentration at the end of the test should be 3.0 mg/L in control and test vessels.

As no immobility or other signs of stress were observed in the control and dissolved oxygen concentration remained at 3 mg/L. This test has satisfied all validity criteria.

Validity criteria fulfilled:
yes
Conclusions:
Under the study conditions, the 48 h EC50 for the test substance in Daphnia was determined to be >100 mg/L (nominal).
Executive summary:

A study was conducted to determine the acute toxicity of the test substance to Daphnia magna according to OECD Guideline 202, in compliance with GLP. Twenty test organisms were exposed to nominal concentrations of 0, 6.25, 12.5, 25, 50 and 100 mg/L for 48 h under static conditions. No confirmatory analysis of the test solutions was performed as a validated analytical method could not be developed. Immobilisation was recorded at 24 and 48 h and compared with control values. Based on immobility compared to the controls (p <0.05), the 48 h NOEC was determined to be 100 mg/L (nominal) and the LOEC was >100 mg/L (nominal). No other symptoms of toxicity were observed. Based on the study results, the 48 h EC50 was calculated to be >100 mg/L (nominal). As no immobility or other signs of stress were observed in the control and dissolved oxygen concentration remained at ≥3 mg/L, all validity criteria were considered to be met. Under the study conditions, the 48 h EC50 for the test substance was determined to be >100 mg/L (nominal) (Scymaris, 2018).

Description of key information

Based on the study results, the 48 h EC50 for the test substance in Daphnia was determined to be >100 mg/L (nominal).

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Effect concentration:
100 mg/L

Additional information

A study was conducted to determine the acute toxicity of the test substance to Daphnia magna according to OECD Guideline 202, in compliance with GLP. Twenty test organisms were exposed to nominal concentrations of 0, 6.25, 12.5, 25, 50 and 100 mg/L for 48 h under static conditions. No confirmatory analysis of the test solutions was performed as a validated analytical method could not be developed. Immobilisation was recorded at 24 and 48 h and compared with control values. Based on immobility compared to the controls (p <0.05), the 48 h NOEC was determined to be 100 mg/L (nominal) and the LOEC was >100 mg/L (nominal). No other symptoms of toxicity were observed. Based on the study results, the 48 h EC50 was calculated to be >100 mg/L (nominal). As no immobility or other signs of stress were observed in the control and dissolved oxygen concentration remained at ≥3 mg/L, all validity criteria were considered to be met. Under the study conditions, the 48 h EC50 for the test substance was determined to be >100 mg/L (nominal) (Scymaris, 2018).

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