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 fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

23 APR 2013 - 27 APR 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

OECD Guideline for Testing ofChemicals, No. 203: "Fish, Acute Toxicity Test" adopted July 17, 1992

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Version / remarks:

Council Regulation (EC) No. 440/2008 laying down test methods pursuant to Regulation (EC) No. 1907/2006 of the European Parliament and the council on the
Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH). C.1. „Fish, Acute Toxicity Test"

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 100 mg/L (nominal) and control
- Sampling method: Samples of the test material concentration and the control were taken at the start ofthe study and after 96 hours. The samples obtained were mixed with Acetonitrile proportionally 1 : 1 and immediately deep frozen (approx:imately at -20°C).
- Sample storage conditions before analysis: All samples were stored at -20 ± 2 °C immediately after sampling. One replicate per concentration was stored at 6 ± 2 °C from 2013-05-14. Prepared samples were stored at room temperature until analysis.

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 membrane filter with a pore size of 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.): The medium was a clear preparation after filtration through a membrane filter with a pore size of 0.2 μm. During the study the test medium changed into a light cloudy/opalescent liquid, what was observed first after 48 hours.

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: zebrafish (Danio rerio)
- Source: The test fish were originally obtained by the "Institut für Gewässerökologie und Binnenfischerei", Berlin, Germany and further bred in the laboratories of Merck KGaA.
- Length at study initiation: 2.0 ± 1.0 cm
- Weight at study initiation: 6.6 - 6.9 g
- Method of breeding: Before start of exposure, the fish were kept at least 12 days in holding medium (reconstituted water) in an air-conditioned room.

ACCLIMATION
- Acclimation period: At least 12 days.
- Acclimation conditions (same as test or not): Before start of exposure, the fish were kept at least 12 days in holding medium (reconstituted water) in an air-conditioned room.
- Feeding frequency during acclimation: The fish were fed every day; the last feeding took place 24 hours before start of the study.
- Health during acclimation (any mortality observed): No fish died during the acclimatization phase. Therefore, the fish were suitable for the present study.

FEEDING DURING TEST
- Frequency: The fish were not fed during the study.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

250 mg/L CaCO3

Test temperature:

19.9 - 21.8 °C
The water temperature was transiently outside the target range of 21 - 25 °C. This minor and short deviation did not influence the integrity or outcome ofthe study.

pH:

7.40 - 7.80

Dissolved oxygen:

71.7 - 98.0 % ASV

Nominal and measured concentrations:

nominal: 100 mg/L
measured initial: 43.4 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: full glass tank
- Type (delete if not applicable): slightly covered by a glass plate
- Material, size, headspace, fill volume: 10 L
- Aeration: The glass tanks were covered in a way to allow air exchange but there was no additional aeration.
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: During the experimental phase a maximum holding density of 1 g fish per liter medium was not exceeded.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Reconstituted water was used as the vehicle. Therefore, demineralized water with conductivity below 5 μS/cm was mixed with analytically pure sa!ts.
Composition:
CaCl2 x 2H2O: 2.0 mmol/L ( = 294.0 mg/L)
MgSO4 X 7H2O: 0.5 mmol/L ( = 123.3 mg/L)
NaHCO3: 0.75 mmol/L ( = 65.8 mg/L)
KCI: 0.075 mmol/L ( = 5.8 mg/L)
Properties:
Hardness of water: 2.5 mmol/L = 250 mg/L as CaCO3
Alkalinity: 0.8 mmol/L
Ratio of Ca:Mg = 4: 1 and Na:K =10:1
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: Lighting was controlled by a timer to provide a 12 hour light and 12 hour dark regime.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): During exposure, the fish were observed in accordance with the laboratory record forms. Evaluation criteria were loss of equilibrium, changes in swimming behavior, respiratory function, pigmentation, etc. as well as the death of the animals. Fish were considered dead, if touching produced no reaction. Dead fish were removed immediately.

TEST CONCENTRATIONS
- Results used to determine the conditions for the definitive study: In a pretest no mortality was observed at a concentration of 100 mg/L under open static conditions.

Reference substance (positive control):

no

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 43.4 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

72 h

Dose descriptor:

LC50

Effect conc.:

> 43.4 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

48 h

Dose descriptor:

LC50

Effect conc.:

> 43.4 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

24 h

Dose descriptor:

LC50

Effect conc.:

> 43.4 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

- Behavioural abnormalities: The fish of the test material group swam on the bottom of the aquarium and showed accelerated respiration starting 24 hours after start of exposure up to 72 hours. The observed clinical signs of toxicity in the surviving fish were completely reversible as they were not observed after 96 hours of exposure anymore.
- Observations on body length and weight: no
- Other biological observations: no
- Mortality of control: no
- Other adverse effects control: No signs of toxicity were seen in the control group.
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: The medium was a clear preparation after filtration through a membrane filter with a pore size of 0.2 μm. During the study the test medium changed into a light cloudy/opalescent liquid, what was observed first after 48 hours.

Validity criteria fulfilled:

yes

Conclusions:

The study was conducted under GLP according to OECD 203 and EU Method C.1. The method is to be considered scientifically reasonable with no deficiencies in documentation or deviations from the guidelines and the validity criteria were met. Hence, the results can be considered as reliable to assess the toxicity of the test substance towards fish.
The test material, solved in reconstituted water, was tested in an open static test system. Under the conditions of the present study, an aqueous solution of 100 mg/L test material caused lethality in one out of 10 exposed fish 48 hours after start of exposure. Other signs of toxicity (swimming on the bottom, accelerated respiration) in the surviving fish were completely reversible. The 96h LC50 based on the initial analytical concentration was > 43.4 mg/L (nominal > 100 mg/L) and, thus, could not be determined in this study.

Executive summary:

This study was conducted according to OECD TG 203 and EU Method C.1 under GLP to investigate the acute toxicity of the test material in the zebrafish (Danio rerio). For this purpose, 10 zebrafish were exposed to a nominal test material concentration of 100 mg/L (limit test) over 96 hours, under defined conditions. Additionally, one control group (10 fish) was used. The fish were observed for signs of toxicity or death for 96 hours.

Stability testing of the test material at a nominal concentration of 100 mg/L in water in open vessels revealed a recovery rate of 96.8 % after 96 hours. Therefore, this study was performed as a static test in open vessels.
Analytical controls of the test material preparation were performed at the start of the study and after 96 hours.

The analytically determined test material concentration immediately after preparation of the medium was 43 % of the nominal concentration and 96 hours thereafter 47 %.
No mortality was observed in the control group. In the test group, 1 zebrafish died 48 hours after start of exposure.
No signs of toxicity were seen in the control group. The fish of the test material group swam on the bottom of the aquarium and showed accelerated respiration starting 24 hours after start of exposure up to 72 hours.
During the experimental phase of the study, the test material concentration could be maintained within ± 20 % of the initial analytical concentration. Therefore, the LC₅₀ values were calculated with the initial analytical concentration.

For the test material, the following analytical LC₅₀ values for zebrafish (Danio rerio) were determined:

24h  LC₅₀ =  43.4 mg/L  (nominal > 100 mg/L)
48h  LC₅₀ =  43.4 mg/L  (nominal > 100 mg/L)
72h  LC₅₀ =  43.4 mg/L  (nominal > 100 mg/L)
96h  LC₅₀ =  43.4 mg/L  (nominal > 100 mg/L)

Description of key information

1) Short-term toxicity to fish: LC₅₀ (96h) > 43.4 mg/L (initial analytical concentration) for Danio rerio based on mortality (static, OECD 203, GLP)

Additional information

1The study by Gado, A. (2013) was conducted according to OECD TG 203 and EU Method C.1 under GLP to investigate the acute toxicity of the test material in the zebrafish (Danio rerio). For this purpose, 10 zebrafish were exposed to a nominal test material concentration of 100 mg/L (limit test) over 96 hours, under defined conditions. Additionally, one control group (10 fish) was used. The fish were observed for signs of toxicity or death for 96 hours.

Stability testing of the test material at a nominal concentration of 100 mg/L in water in open vessels revealed a recovery rate of 96.8 % after 96 hours. Therefore, this study was performed as a static test in open vessels.
Analytical controls of the test material preparation were performed at the start of the study and after 96 hours.

The analytically determined test material concentration immediately after preparation of the medium was 43 % of the nominal concentration and 96 hours thereafter 47 %.
No mortality was observed in the control group. In the test group, 1 zebrafish died 48 hours after start of exposure.
No signs of toxicity were seen in the control group. The fish of the test material group swam on the bottom of the aquarium and showed accelerated respiration starting 24 hours after start of exposure up to 72 hours.
During the experimental phase of the study, the test material concentration could be maintained within ± 20 % of the initial analytical concentration. Therefore, the LC₅₀ values were calculated with the initial analytical concentration.

For the test material, the following analytical LC₅₀ values for zebrafish (Danio rerio) were determined:

24h  LC₅₀ =  43.4 mg/L  (nominal > 100 mg/L)
48h  LC₅₀ =  43.4 mg/L  (nominal > 100 mg/L)
72h  LC₅₀ =  43.4 mg/L  (nominal > 100 mg/L)
96h  LC₅₀ =  43.4 mg/L  (nominal > 100 mg/L)