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Ecotoxicological information

Short-term toxicity to aquatic invertebrates

Administrative data

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

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018
Report Date:
2018

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
Samples for possible analysis were taken from all test concentrations and the control according to the schedule as follows:

Frequency: At 0 and 48 hours (start and end of test)
Volume: 3 mL from the approximate centre of the test vessels
Storage: Samples were stored in a freezer (≤-15°C) until analysis at the analytical laboratory of the Test Facility

At the end of the exposure period, the replicates were pooled at each concentration before sampling. Additionally, reserve samples of 3 mL were taken for possible analysis. If not used, these samples were stored in a freezer (≤-15°C).

Test solutions

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION:
The batch of the test item was a clear yellow viscous liquid UVCB and not completely soluble in test medium at the loading rates initially prepared. No correction was made for the purity/composition of the test item. Preparation of test solutions was performed under dimmed light conditions with glassware wrapped in aluminium foil to minimize exposure to light.

Preparation of test solutions started with loading rates individually prepared at 1.0, 10 and 100 mg/L. A 31-minute period of ultrasonic waves followed by an overnight period of magnetic stirring was applied to ensure maximum dissolution of the test item in test medium. The obtained mixtures were centrifuged at 10000 rpm for 10 minutes. Subsequently, the supernatant was drained from the pelleted undissolved material using a pipette and the obtained Water Accommodated Fractions (WAFs) were used as test concentrations. All test solutions were clear and colorless at the end of the preparation procedure.

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia magna (crustacea, Cladocera)
- Strain/clone: Straus 1820
- Source: In-house laboratory culture with a known history
- Age of parental stock (mean and range, SD): 2 to 4 weeks
- Feeding during test: None

ACCLIMATION
- Acclimation period: Parental cultures up to 4 weeks old
- Acclimation conditions (same as test or not): Same as test
- Type and amount of food: Fresh water algae
- Feeding frequency: Daily
- Health during acclimation (any mortality observed): Only Daphnids from healthy stock used (2nd to 5th brood, showing no signs of stres such as mortality > 20%, presence of males, ephippia or discoloured animals and no delay in the production of the first brood).

METHOD FOR PREPARATION AND COLLECTION OF EARLY INSTARS OR OTHER LIFE STAGES: Daphnia less than 24 hours old.

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h

Test conditions

Hardness:
180 mg/L as CaCO3
Test temperature:
Varied between 19 and 21°C during the test constant within +/- 1°C.
pH:
Remained within pH 6-9 and did not vary by more than 1.5 units.
Dissolved oxygen:
Dissolved oxygen >/= 3 mg/L at the end of the test
Salinity:
Not specified
Conductivity:
Not specified
Nominal and measured concentrations:
Nominal: Control, 1, 10 and 100 mg/L (loading rates)
Measured: 0 and 46 mg/L (at a loading rate of 100 mg/L and not measured at loading rates of 1 or 10 mg/L)
Details on test conditions:
TEST SYSTEM
- Test vessel: All-glass vessels
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: 50 mL test/control solutions in 60 mL vessels
- Aeration: No
- No. of organisms per vessel: 5 per vessel
- No. of vessels per concentration (replicates): 100 mg/L; 20 per test group and 1 and 10 mg/L; 10 per test group.
- No. of vessels per control (replicates): 4
- No. of vessels per vehicle control (replicates): NA
- Biomass loading rate: 10 mL per Daphnid

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Analytical grade salts were added to tap water purified by reverse osmosis. The culture medium was M7 as per the OECD guideline)

OTHER TEST CONDITIONS
- Adjustment of pH: Not performed
- Photoperiod: Test conducted in darkness
- Light intensity: Not applicable

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Immobility measured at 24 and 48 hours. Immobile are those animals not able to swim within 15 seconds after gentle agitation of the test vessel, even if they can still move their antennae.

VEHICLE CONTROL PERFORMED: No

RANGE-FINDING STUDY
- Test concentrations: Combined range finder/limit test performed at 1, 10 and 100 mg/L loading rates
Reference substance (positive control):
yes
Remarks:
Potassium dichromate

Results and discussion

Effect concentrationsopen allclose all
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat. (dissolved fraction)
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 46 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat. (dissolved fraction)
Basis for effect:
mobility
Details on results:
Measured concentrations:
Samples taken from the highest test concentration were analysed. The measured concentration was 46 mg/L at the start of the test. During the exposure period, the concentration remained stable, i.e. was at 112% relative to initial at the end of the test. Based on these results, effect parameters were expressed as the initially measured concentration and the loading rate. This measured concentration was determined to be the maximum solubility of the potentially soluble constituents of the UVCB substance. As such the EC50 values exceeded the limit of solubility.

Immobility:
No immobility was observed in the control and at the two lowest test concentrations throughout the exposure period. At the end of the test, 30% immobility was observed at the highest concentration tested.
Results with reference substance (positive control):
- Results with reference substance valid? Yes
- Relevant effect levels: 48 hour EC50 was 0.59 mg/L (with the historical range of 0.53 to 0.72 mg/L).
- Limit test: No
- Dose-response test: Yes (concentrations of 0.1, 0.18, 0.32, 0.56, 1.0 and 1.8 mg/L).
- ECx: EC50
Reported statistics and error estimates:
Not performed as EC50 greater than the highest concentration/loading rate tested.

Any other information on results incl. tables

Number of introduced Daphnids and incidence of immobility in the test:

Time (h)

Replicate

Test substance
Loading rate (mg/L)

Control

1.0

10

100

0

A

5

5

5

5

B

5

5

5

5

C

5

 -

 -

5

D

5

 -

 -

5

Total introduced

20

10

10

20

24

A

0

0

0

2

B

0

0

0

1

C

0

-

 -

0

D

0

 -

0

Total immobilised

0

0

0

3

Effect %

0

0

0

15

 

 

 

 

48

A

0

0

0

2

B

0

0

0

1

C

0

 -

 -

1

D

0

 -

 -

2#

Total immobilised

0

0

0

6

Effect %

0

0

0

30

Test sample analysis:

Time of sampling
[hours]

Date of sampling

Date of
analysis1

Loading rate2

[mg/L]

Analyzed concentration
[mg/L]

Relative to
initial
[%]

0

19 Jun 2018

27 Jun 2018

0

n.d.

 

 

 

 

100

45.6

 

48

21 Jun 2018

27 Jun 2018

0

n.d.

n.a.

 

 

 

100

51.1

112

1 Samples were stored in the freezer (≤ -15°C) until the day of analysis.

2 A water accommodated fraction (WAF) prepared at the loading rate.

n.d. Not detected.

n.a. Not applicable.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
In conclusion the 48 hour EL50 for Daphnia magna exposed to the test substance was greater than the highest loading rate tested (100 mg/L). Based on analytical measurements conducted of dissolved constituents the EC50 was greater than 46 mg/L (initial measured concentration). This was determined to be the the limit of solubility for the dissolved fraction.