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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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
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
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
yes
Details on sampling:
- Concentrations: nominal 4.27, 9.39, 20.7, 45.5, and 100 mg/L and control
- Sampling intervals: start and end of the renewal periods
- Sampling method: Samples from expired media were taken from the pooled media from replicate vessels.
- Sample storage conditions before analysis: no storage prior to analysis
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: water accommodated fractions (WAFs)
At each concentration, the test substance was dispersed in dilution medium in an aspirator. The bulk preparations (5 L) were stirred for approximately 24 hours in the dark, after which a syphon was added to the mid-vessel portion of each aspirator and the preparations were then left to stand for a further approximate 24 hours in the dark. An aliquot (1 L) of the mid-vessel portion was siphoned from each preparation vessel and discarded, after which a further 1 L was removed mid-vessel to provide the Water Accommodated Fractions (WAFs).
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Source: in-house culture originating from a strain supplied by the National Institute for Applied Chemical Research (IRCHA), France
- Age at study initiation: < 24 h
- Method of breeding: Stock cultures of Daphnia magna were maintained in glass vessels containing approximately 0.5 to 0.8 litres of Elendt M4 culture medium in a temperature-controlled laboratory at nominally 20 ± 2°C. A photoperiod of 16 hours light : 8 hours dark was maintained, with periods of subdued lighting at the beginning and end of each light phase. The culture medium was renewed three times each week.
Cultures were fed daily with a suspension of the unicellular green algae, Pseudokirchneriella subcapitata, to provide nominally 0.1 to 0.2 mg carbon per daphnid, per day, except during the initial two days when a slightly lower ration was given. Culture conditions ensured that the stock animals reproduced by parthenogenesis.
The day before the start of the study, all juvenile Daphnia were removed from the laboratory cultures. The following morning, juveniles produced by the gravid (egg-bearing) adult Daphnia were removed from the culture vessels, held in a separate holding vessel and used in the test.
- Feeding during test: none

ACCLIMATION
- Acclimation period: not relevant, conditions were the same for maintenance and testing
Test type:
semi-static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
48 h
Hardness:
267 and 251 mg/L as CaCO3 for the two batches of Elendt M4 medium used
Test temperature:
20.9 ± 0.3 °C
pH:
7.7-8.1
Dissolved oxygen:
88-96% air saturation value (>3 mg/L)
Nominal and measured concentrations:
nominal loading rates: 4.27, 9.39, 20.7, 45.5 and 100 mg test material/L
At all concentrations, the levels of carbon ranged between 0.00 and 1.17 mg C/L after background correction.
The levels of carbon measured in the test media at 4.27 to 20.7 mg/L were at or below the limit of detection of the assay system (1 mg C/L), exposure concentration at these levels could not be accurately quantified. At 45.5 and 100 mg/L, the overall geometric mean measured TOC levels were 0.573 and 0.952 mg C/L, respectively.
Details on test conditions:
TEST SYSTEM
- Test vessel: covered glass jars containing 100 mL of medium
- Aeration: none
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- Biomass loading rate: 20 mL medium per organism

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: deionised, reverse osmosis water
- Total organic carbon: 0.9-1.2 mg C/L
- Alkalinity: 58 and 39 mg /L as CaCO3 for the two batches of Elendt M4 medium used
- Ca/Mg ratio: 3.7:1
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 hours light : 8 hours dark with periods of subdued lighting at the beginning and end of each light phase

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : numbers of mobile, immobile and floating Daphnia counted approx. 24 and 48 hours after the start of the study

RANGE FINDING STUDY
- Test concentrations: nominal loading rates of 1, 10 and 100 mg/L
- Results used to determine the conditions for the definitive study: 10% immobility at 10 and 100 mg/L after 48 hours. Not considered significant.
Reference substance (positive control):
yes
Remarks:
potassium dichromate
Key result
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: results are expressed in terms of loading rates
Duration:
48 h
Dose descriptor:
NOELR
Effect conc.:
>= 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: results are expressed in terms of loading rates
Details on results:
- Behavioural abnormalities: none
- Other biological observations: After 48 hours, 10% immobilisation occurred at a nominal loading rate of 100 mg/L, which was not considered significant; no immobility occurred at any other test concentration (for details see Table 1: Cumulative immobilisation).
- Mortality of control: none
- Other adverse effects control: none
- Abnormal responses: none

At all concentrations, the test media were colourless. In expired medium, particulate matter was observed on the surface of test vessels at nominal loading rates of 20.7 to 100 mg/L.
Results with reference substance (positive control):
- 48-hour EC50: 1.11 mg/L (typical range obtained in the performing laboratory: 1.1-1.7 mg/L)
Reported statistics and error estimates:
The “no observed effect loading rate” (NOELR) was derived by direct inspection of the data on the immobility of the animals. An incidence rate of more than 10% was considered to be significant.

Table 1: Cumulative immobilisation

 

Nominal loading rate (mg/L)

Cumulative numbers of immobile Daphnia

24 hours

48 hours

R1

R2

R3

R4

total

%

R1

R2

R3

R4

total

%

Control

0

0

0

0

0

0

0

0

0

0

0

0

4.27

0

0

0

0

0

0

0

0

0

0

0

0

9.39

0

0

0

0

0

0

0

0

0

0

0

0

20.7

0

0

0

0

0

0

0

0

0

0

0

0

45.5

0

0

0

0

0

0

0

0

0

0

0

0

100

0

0

0

1

1

5

0

0

1

1

2

10

R: replicate number

 

Validity criteria fulfilled:
yes
Remarks:
The number of floating and/or immobile Daphnia in the control group did not exceed 10% and the concentration of dissolved oxygen was > 3 mg/L (≥60% of the air saturation value at the temperature used) at the end of the test.

Description of key information

48-h EC50 (based on loading rates) >100 mg/L for Daphnia magna (OECD TG 202, EU C.2)

Key value for chemical safety assessment

Additional information

The results indicate that the test material is not toxic to daphnia up to the limits of its water solubility (<1 mg/L) and a NOEC and thus a PNEC cannot be determined.