[No public or meaningful name is available]

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

April 26, 2022 to August 09, 2022

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

conducted under GLP conditions

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

2004

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Remarks:

GC-MS

Details on sampling:

Samples of test solutions were taken to determine the actual test item concentrations in comparison to the nominally applied concentrations.
Control and test solutions were sampled in duplicate. The duplicate samples were kept separately as a reserve. The volume of each sample was recorded.
Samples of a defined volume (20 mL) were taken from the designated test and/or stock solutions. After sampling and before shipment, all samples were stored in amber glass bottles in the dark at a temperature of ≤ -18°C if not advised otherwise. A record was kept for each sample.
Samples were transferred to the test site for chemical analysis (address see section 1.3 of the final report) under the required storage conditions. The dates of transfer of the samples from ECT to the laboratory for chemical analysis were recorded in the raw data.
Analysis was performed according to an analytical method validated by the test site for chemical analysis. Further details describing the analytical procedures are contained in the analytical phase report attached.

Vehicle:

yes

Details on test solutions:

Based on the results of a preceding non-GLP range finding acute toxicity test to Daphnia magna, the following nominal concentrations in a geometrical series (spacing factor: 2.2) were tested in the definitive test:
0.342, 0.751, 1.65, 3.64 and 8.00 mg test item/L.
The test period was 48 hours; exposure of test organisms to the test solutions and test medium in a semi-static system with one renewal after 24 hours.
Four replicates were used per test item concentration and for the controls. Each replicate vessel contained five daphnids.

Two stock solutions (S1) of the test item were prepared at start and on day one (24 hours) of the study. Therefore, 0.01593 or 0.01631 g were dissolved in 2 L M4 medium using a 2 L measuring flask respectively, resulting in a nominal concentration of 8.0 mg test item/L. The test item was rinsed into the flask and was filled-up to the mark. The stock solutions were stirred on a magnetic stirrer for 20‒25 minutes at 750 rpm at ambient temperature in the dark. The stock solution was left for 15 minutes without stirring and no undissolved test item was
observed in the stock solutions. These stock solutions were used to prepare the final test solution.

Test organisms (species):

Daphnia magna

Details on test organisms:

Species: The test system used in this study was Daphnia magna STRAUS (clone M10), a water flea, cultured at ECT Oekotoxikologie GmbH since December 22, 2011. The selection of the test system is based on the requirements of the test guideline.
Origin: The organisms were originally supplied by KU Leuven, Belgium.
The health of the Daphnia culture and the overall quality of the test organisms is monitored regularly by using the reference toxicant potassium dichromate (K2Cr2O7) in an acute test design in order to determine the sensitivity of the organisms from the culture and to
demonstrate that the test conditions are appropriate.
The most recent reference test with potassium dichromate performed in a separate study (Study No. IDA2203; March 2022) resulted in an EC50(24 h) of 1.528 mg/L.
According to the OECD guideline No. 202 (OECD, April 2004) EC50(24 h)-values for potassium dichromate obtained from different laboratories should range between 0.6 mg/L and 2.1 mg/L.
The toxicity of the reference item is within this range. Therefore, the results of this reference test are acceptable and the test conditions are reliable and the sensitivity of the test system could be demonstrated.


The culture conditions of Daphnia magna as summarised below are similar to the test conditions.
Material of stock vessel: Glass
Volume of medium per stock vessel: 1.8 L
Depth of medium in the stock vessel: Approximately 14.5 cm
Number of daphnids kept as stock per culture vessel (batch size): Approximately 20
Separation of adult and young daphnids: 2–3 times per week
Medium: Elendt medium M4
Renewal of medium: Twice per week
Temperature: Within required ranges (202 °C)
Photoperiod (light/dark): 16/8 h
Light intensity: Within required ranges (50–1000 lx)
Aeration: None
Food: Algae (live Desmodesmus subspicatus), supplemented by instant baker’s yeast suspension and TetraMin® suspension
Feeding frequency: 3–4 times per week

The selected holding vessels designated for production of the neonates for the test were cleared of all mixed-aged young daphnids on the day before test start. These vessels had a record of more than one brood and contained parental daphnids aged less than 35 days. On the day of test start (day 0) all newly produced neonates of these vessels were < 24 hours old.
These neonates were collected for the test.

Test type:

semi-static

Water media type:

freshwater

Remarks:

Medium: Elendt M4

Limit test:

no

Total exposure duration:

48 h

Hardness:

252 mg/L CaCO3

Test temperature:

20.1-21.0°C

pH:

7.6-7.8

Dissolved oxygen:

8.8-9.5 O2/L

Conductivity:

660 microS/cm

Nominal and measured concentrations:

Nominal concentrations: 0.342, 0.751, 1.65, 3.64 and 8.00 mg test item/L and a control.

Reference substance (positive control):

yes

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

1.08 mg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

1.87 mg/L

95% CI:

>= 1.44 - <= 2.45

Nominal / measured:

meas. (TWA)

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

0.52 mg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

The measured test item concentrations were not stable during the test period, and ranged between 15.4% and 128% of the nominal concentrations within each renewal period of 24 hours. To evaluate the biological results, the time-weighted-mean measured concentration was calculated for each treatment level throughout the exposure period. The following time weighted-mean concentrations were determined: 0.246, 0.529, 1.08, 3.17 and 7.40 mg test item/L.
The biological endpoints are therefore expressed based on nominal and time-weighted mean concentrations.

Results with reference substance (positive control):

The health of the Daphnia culture and the overall quality of the test organisms is monitored regularly by using the reference toxicant potassium dichromate (K2Cr2O7) in an acute test design in order to determine the sensitivity of the organisms from the culture and to demonstrate that the test conditions are appropriate.
The most recent reference test with potassium dichromate performed in a separate study (Study No. IDA2203; March 2022) resulted in an EC50(24 h) of 1.528 mg/L.
According to the OECD guideline No. 202 (OECD, April 2004) EC50(24 h)-values for potassium dichromate obtained from different laboratories should range between 0.6 mg/L and 2.1 mg/L.
The toxicity of the reference item is within this range. Therefore, the results of this reference test are acceptable and the test conditions are reliable and the sensitivity of the test system could be demonstrated.

Validity criteria fulfilled:

yes

Conclusions:

The 48 hour EC50 for Daphnia magna exposed to the test substance was determined to be 1.87 mg/L based on time-weighted mean measured concentrations.

Executive summary:

The aim of the study was to determine the acute toxicity of the test item to the water flea Daphnia magna according to the OECD guideline No 202.

The 48 hour EC50 for Daphnia magna exposed to the test substance was determined to be 1.87 mg/L.

Description of key information

The aim of the study was to determine the acute toxicity of the test item to the water flea Daphnia magna according to the OECD guideline No 202.

The 48 hour EC50 for Daphnia magna exposed to the test susbtance was determined to be 1.87 mg/L based on time-weighted-mean measured concentrations.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Dose descriptor:

EC50

Effect concentration:

1.87 mg/L