Sodium 6,14-diethyl-1,1,1,2,2,18,18,19,19,19-decafluoro-8,12-dioxo-10-({[1-(2,2,3,3,3-pentafluoropropoxy)butan-2-yl]oxy}carbonyl)-4,7,13,16-tetraoxanonadecane-9-sulfonate

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:

10 DEC 2012 - 05 JUL 2013

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)

Version / remarks:

OECD Guideline for Testing of Chemicals, No. 202: "Daphnia sp. Acute Immobilization Test and Reproduction Test", 2004.

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

Council Regulation (EC) No. 440/2008 laying down test methods persuant to Regulation (EC) No. 1907/2006 of the European Parliament and the council on the
Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH). C.2 ,,Acute toxicity for Daphnia"

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0 and 100 mg/L (nominal)
- Sampling method: Samples of the test material concentration and the control were taken at the start of the study and after 48 hours.
- Sample storage conditions before analysis: The samples obtained were mixed with Acetonitrile proportionally 1: 1 and immediately deep frozen (approximately at -20°C).

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The test medium (reconstituted water and test material) was prepared freshly. Therefore, the calibrated flask with test medium was treated in an ultrasonic device for 1 hour. Subsequently, the preparation was stirred with a magnetic stirrer for further 23 hours. Afterwards, the formulation was passed through a single use syringe filter (pore size 0.2 µm). The filtrate was used for the study.
- Controls: For the control, reconstituted water without any addition of solvent or test material was used.
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): No remarkable observations were made concerning the appearance of the solution of the test material preparation. The test medium was a clear preparation and stayed unchanged throughout the study

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Strain/clone: Daphnia magna Straus
- Age at study initiation (mean and range, SD): When the study was planned, all offspring were separated from their parents less than 24 hours before start of the experimental part, so on the following morning when exposure began, all offspring were not older than 24 hours.
- Stage and instar at study initiation: neonates
- Method of breeding: The clone was bred in the laboratories of Merck KGaA at a water temperature of 20 ± 2°C controlled by MCPS (Multi Channel Process System) in a separate vessel. To avoid an acclimation phase before start of the study, reconstituted water was used during culture as well as during toxicity studies.The daphnids were kept in individual 100 mL test vessels containing approximately 60 mL reconstituted water. They were fed with green algae whenever necessary, but at least once a week. During the breeding phase, outside of studies, the offspring were separated from their parents during the change of reconstituted water.
- Source: Daphnia magna Straus of the parent generation was originally bred by IBACON GmbH (Rossdorf, Gerrnany).
- Age of parental stock (mean and range, SD): Parent animals were used for reproduction until they were about 6 weeks old. Then they were replaced by newborn offspring.
- Feeding during test : The daphnids were not fed during the experimental phase.

METHOD FOR PREPARATION AND COLLECTION OF EARLY INSTARS OR OTHER LIFE STAGES, INCLUDING CULTURING CONDITIONS:
Before start of the experimental phase, the young daphnids were separated from their different parent animals and put into a collective test vessel containing reconstituted water. The daphnids were visually inspected for motility. If daphnids did not make any swimming movements within 15 seconds after their containers have been gently agitated, they were considered to be immobile and were not be used for the study. At the start of the experimental phase they were cautiously removed using a pipette, separated from the reconstituted water using a fine-mesh sieve, and placed into the test vessels containing test media (test material group) or reconstituted water (control).

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Hardness:

about 250 mg/L CaCO3

Test temperature:

21 °C

pH:

7.89 - 7.96

Dissolved oxygen:

8.07 - 8.17 mg/L

Nominal and measured concentrations:

nominal: 100 mg/L
measured initial: 27.8 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: glass vessels
- Type (delete if not applicable): open
- Volume of solution: Test vessels were filled with at least 20 mL either reconstituted water (control) or test media (test material group).
- Aeration: The control medium and test medium were not aerated during the experimental phase.
- 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: The water was composed according to ELENDT (1990). The reconstituted water contained the following chemicals (analytical grade), which were dissolved in fully demineralized water to obtain the following concentrations:
Main components
CaCl2 * 2H2O: 293.80
MgSO4 * 7H2O: 123.30
NaHCO3: 64.80
KCl: 5.80
Na2SiO3 * 9H2O: 10.00
NaNO3: 0.27
KH2PO4: 0.14
K2HPO4: 0.18

Trace elements
B: 0.5000
Fe: 0.2000
Mn: 0.1000
Li, Rb, and Sr: 0.0500
Mo: 0.0250
Br: 0.0125
Cu and Zn: 0.0063
Co and I: 0.0025
Se: 0.0010
V: 0.0003
Na2EDTA * 2H2O: 2.5000

Vitamins
Thiamin: 0.07500
B12: 0.00100
Biotin: 0.00075

After preparation the reconstituted water was aerated for 24 hours. Hardness: about 2.5 mmol/L (about 250 mg/L CaCO3), pH: 7.9 ± 0.3, after an aeration of 24 hours.

- Culture medium different from test medium: To avoid an acclimation phase before start of the study, reconstituted water was used during culture as well as during toxicity studies.

OTHER TEST CONDITIONS
- Adjustment of pH: not specified
- Photoperiod: Lighting was controlled by a timer to provide a 16 hour light and 8 hour dark regime.
- Light intensity: The mean light intensity measured immediately before start of the study was 743 Lux and 745 Lux at the end ofthe study.
- Other: The study was performed in an air-conditioned room. The vessels were placed on a dark surface in a plastic box with a lid.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : 24 and 48 hours after the daphnids had been placed into the reconstituted water (control) or respective test medium (test material group ), they were visually examined for their ability to swim. Those daphnids that did not make any swimming movements within 15 seconds after the test vessel had been gently agitated, were regarded as not being able to swim.

RANGE-FINDING STUDY
- Test concentrations: 100 mg/L (nominal)
- Results used to determine the conditions for the definitive study: No immobilization was observed in the pre-test at the concentration of nominal 100 mg/L under open static conditions.

Reference substance (positive control):

yes

Remarks:

A reference study with potassium dichromate as positive control was performed at the test facility from 31.07. to 02.08.2012 to check the sensitivity of the test system as recommended by guidelines (Study-No.: T18392).

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

> 27.8 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 27.8 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Behavioural abnormalities: none
- Mortality of control: no
- Other adverse effects control: no
- Immobilisation of control: no
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No remarkable observations were made concerning the appearance of the solution of the test material preparation. The test medium was a clear preparation and stayed unchanged throughout the study

Results with reference substance (positive control):

- Results with reference substance valid? yes
- Relevant effect levels: Under the given experimental conditions potassium dichromate showed a 24h EC50 value which was within the range of the published data of 0.6 to 1. 7 mg/L (Council Regulation (EC) No. 440/2008) and 0.6 to 2.1 mg/L (OECD Guideline No. 202).
- Dose-response test: Test material concentrations of 0.31, 0.46, 0.69, 1.02, and 1.52 mg/L were used in this study.
- ECx: EC50 (24h) = 1.018 mg/L (95% confidence interval: 0.887 - 1.195 mg/L), EC50 (48h) = 0.769 mg/L (95% cinfidence interval: 0.672 - 0.883 mg/L)

Validity criteria fulfilled:

yes

Conclusions:

Under the conditions of the present study, an aqueous solution of 100 mg/L of the test material revealed no aquatic toxicity in the test system.
The 48h EC50 was > 27.8 mg/L (> 100 mg/L, nominal) and could thus not be determined in this study.
The study was conducted under GLP according to OECD Guideline 202 and EU Method C.2 on the registered substance itself. The method is to be considered scientifically reasonable with no deficiencies in documentation or deviations from the guidelines, the validity criteria were met. Hence, the results can be considered as reliable to assess the toxicity of the test substance towards daphnids.
Daphnia magna were exposed to an aqueous solution from a test material concentration of nominal 0.1 g/L (limit-test) over 48 hours in an open system under defined conditions. After an exposure to a saturated aqueous solution of a nominal concentration of 0.1 g/L for 48 hours the test animals were not affected. As the analysis of a saturated aqueous solution revealed that the water solubility of the test material was < 0.01 g/L and the limit of quantification of the analytical method was described to be about 0.01 g/L, the test material concentrations in the aqueous medium at the start and the end of the study could not be quantified.
Based on the obtained biological results the 24h and 48h EC50 as well as the 24h and 48h EC10 for Daphnia magna were determined to be > 0.1 g/L, respectively.

Executive summary:

This study was performed according to GLP and the methods applied are fully compliant with OECD TG 202.

The objective of this study was to evaluate the influence of the test material on the immobilization of Daphnia magna.

For this purpose, juvenile daphnids were exposed to an aqueous test material solution over 48 hours, under defined conditions. The daphnids were observed for immobilization 24 and 48 hours after placing in the test medium. The study comprised of four test vessels in the test material group containing five daphnids each, i.e. 20 daphnids per concentration. Additionally, one control group (20 daphnids) was used.

Daphnids were exposed to a nominal test material concentration of 100 mg/L (limit test). A stability test of the test material at a nominal concentration of  100 mg/L in reconstituted water in open vessels revealed a recovery rate of 89.9% after 48 hours. Therefore, the test was performed as a static test in open vessels.

After exposure to the nominal test material concentration of 100 mg/L the daphnids were not affected.

Analytical controls of the test material preparation were performed at the start of the study and after 48 hours.

The analytically determined test  material concentration immediately after  preparation of the medium was 28 % and 48 hours thereafter 25 % of the nominal concentration.

During the experimental phase of the study, the test material concentration could be maintained at levels of > 80 % of the initialy measured concentration. Therefore, the EC₅₀ values were calculated with the measured initial concentration.

For the test material, the following nominal EC₅₀ values for daphnids were determined:

24h EC₅₀ > 27.8 mg/L (> 100 mg/L, nominal)

48h EC₅₀ > 27.8 mg/L (> 100 mg/L, nominal)

Description of key information

Short-term toxicity to aquatic invertebrates: EC₁₀ (48h) and EC₅₀ (48h) > 27.8 mg/L (initial analytical concentration) for Daphnia magna based on immobilisation (static, OECD 202, GLP)

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

27.8 mg/L

Additional information

This study was performed according to GLP and the methods applied are fully compliant with OECD TG 202.

The objective of this study was to evaluate the influence of the test material on the immobilization of Daphnia magna.

For this purpose, juvenile daphnids were exposed to  an aqueous test  material solution over 48 hours, under defined conditions. The daphnids were observed for immobilization 24 and 48 hours after placing in the test medium. The study comprised of four test vessels in the test material group containing five daphnids each, i.e. 20 daphnids per concentration. Additionally, one control group (20 daphnids) was used.

Daphnids were exposed to a nominal test material concentration of 100 mg/L (limit test). A stability test of the test material at a nominal concentration of  100 mg/L in reconstituted water in open vessels revealed a recovery rate of 89.9% after 48 hours. Therefore, the test was performed as a static test in open vessels.

After exposure to the nominal test material concentration of 100 mg/L the daphnids were not affected.

Analytical controls of the test material preparation were performed at the start of the study and after 48 hours.

The analytically determined test  material concentration immediately after  preparation of the medium was 28 % and 48 hours thereafter 25 % of the nominal concentration.

During the experimental phase of the study, the test material concentration could be maintained at levels of > 80 % of the initialy measured concentration. Therefore, the EC₅₀ values were calculated with the measured initial concentration.

For the test material, the following nominal EC₅₀ values for daphnids were determined:

24h EC₅₀ > 27.8 mg/L (> 100 mg/L, nominal)

48h EC₅₀ > 27.8 mg/L (> 100 mg/L, nominal)