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Ecotoxicological information

Short-term toxicity to fish

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Reference
Endpoint:
short-term toxicity to fish
Type of information:
experimental study
Adequacy of study:
key study
Study period:
17 December 2014 to 21 January 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 203 (Fish, Acute Toxicity Test)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.1 (Acute Toxicity for Fish)
Deviations:
no
Qualifier:
according to
Guideline:
other: OECD 23 Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
Directive 2004/9/EC
Specific details on test material used for the study:
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature in the dark
Analytical monitoring:
yes
Details on sampling:
- Concentrations: Prepared as WAF, tested as a limit test (100 mg/L loading rate)
- Sampling method: Water samples were taken from the control and the 100 mg/L loading rate WAF vessels at 0 and 72 hours from fresh median and at 24 and 96 hours from old media. Duplicate samples were taken for quantitative analysis.
- Sample storage conditions before analysis: Samples were frozen prior to analysis
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: 2200 mg of the test material was added to the surface of 22 litres of test water to give a 100 mg/L loading rate. The test water was stirred by magnetic stirrer using a stirring rate such that a vortex was formed to give a dimple at the water surface. The stirring was stopped after 23 hours and the mixture allowed to standard for 1 hour. A wide bore glass tube covered at one end with Nescofilm was submerged into the vessel, sealed end down to a depth of approximately 4 cm from the bottom. A length of Tygon tubin was inserted into the glass tube and pushed through the seal. The aqueous phase was removed by mid-depth siphoning (the first 75-100 mL discarded) to give the 100 mg/L loading rate WAF. The vortex depth was recorded at the start and end of the mixing period.
- Controls: Untreated
- Evidence of undissolved material: Microscopic inspection of the WAF showed no micro-dispersion or undissolved test material.
Test organisms (species):
Oncorhynchus mykiss (previous name: Salmo gairdneri)
Details on test organisms:
TEST ORGANISM
- Common name: Rainbow Trout
- Age at study initiation : Juvenile fish
- Length at study initiation : mean standard length 4.5 cm (SD = 0.44)
- Weight at study initiation: mean weight 1.18 g (SD 0.31)

ACCLIMATION
- Acclimation period: 7 days
- Acclimation conditions: Acclimated to test conditions
- Type and amount of food during acclimation: Commercial trout pellets
- Health during acclimation (any mortality observed): 7 mortalities (2.6 % of stock population) in the 7 days prior to the start of the test

FEEDING DURING TEST
- Frequency: Discontinued 24 hours prior to test initiation.
Test type:
semi-static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
96 h
Hardness:
140 mg/L as CaCO3
Test temperature:
14-15 °C
pH:
7.6-8.5
Dissolved oxygen:
9.6 mg O2/L
Conductivity:
Average 411.711 µS/cm at 20 °C
Nominal and measured concentrations:
WAF 100 mg/L Loading Rate
Details on test conditions:
TEST SYSTEM
- Test vessel: Glass exposure vessels
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: 25-30 litre glass vessels containing 20 litres of test media.
- Aeration: Aerated via narrow bore glass tubes
- Renewal rate of test solution (frequency/flow rate): Daily renewal of test media
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.41 bodyweight/litre

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Laboratory tap water dechlorinated by passage through an activated carbon filter and partly softened.
- Total organic carbon: 0.996 mg/L (average prior to dechlorination)
- Metals:
Aluminium <9.737 µg/L (average prior to dechlorination)
Antimony 0.465 µg/L (average prior to dechlorination)
Arsenic <0.371 µg/L (average prior to dechlorination)
Cadmium 0.161 µg/L (average prior to dechlorination)
Chromium <0.700 µg/L (average prior to dechlorination)
Lead <2.163 µg/L (average prior to dechlorination)
Copper 0.074 µg/L (average prior to dechlorination)
Iron <25.276 µg/L (average prior to dechlorination)
Manganese <2.561 µg/L (average prior to dechlorination)
Mercury <0.012 µg/L (average prior to dechlorination)
Nickel 4.338 µg/L (average prior to dechlorination)
Selenium <0.745 µg/L (average prior to dechlorination)
Sodium 22 µg/L (average prior to dechlorination)
- Pesticides: Total pesticides 0.007 µg/L (average prior to dechlorination)
- Chlorine: Total chlorine 0.248 mg/L (average prior to dechlorination)
- Culture medium different from test medium: Same water used

OTHER TEST CONDITIONS
- Photoperiod: 16 hours light 8 hours dark with a 20 minute dusk period.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Mortalities and sub lethal effects were recorded at 3, 6, 24, 48, 72 and 96 hours. Death was recorded as the absence of both respiratory movement and response to physical stimulation.
The water temperature, pH and dissolved oxygen concentrations were recorded daily throughout the test. The measurement at 0, 24, 48 and 72 hours represent the freshly prepared test solutions. The measurements taken each test media renewal and at test termination were 24 hour old solutions.

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 1, 10, 100 mg/L
- Results used to determine the conditions for the definitive study: No mortality or sublethal effects were noted during the range-finding study (3 fish observed over 96 hours)
Reference substance (positive control):
no
Key result
Duration:
96 h
Dose descriptor:
LL50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Key result
Duration:
96 h
Dose descriptor:
NOELR
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Details on results:
- Behavioural abnormalities: No sublethal effects observed
- Mortality of control: No mortality observed in the control
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: Observation on the test media were carried out during the mixing and testing of the WAFs.
At the start of the mixing period for the 100 mg/L loading rate, the test solution was observed to be clear colourless water column with an oily layer of the test material at the surface of the water.
After 23 hours stirring and a 1 hour standing period, the 100 mg/L loading rate was observed microscopically. Microscopic inspection of the WAF showed no micro-dispersions or undissolved test material to be present. After siphoning and for the duration of the test, the test solution was observed to be a clear colourless solution.
Chemical analysis of the test preparations at 0,24, 72 and 96 hours showed measured test material concentrations of less than the LOQ of the analytical method (0.00057 mg/L). This does not infer that no test material was in solution just that any dissolved test material was at a concentration of less than the LOQ.
Reported statistics and error estimates:
An estimate of the LL50 values was given by inspection of the mortality data.

Table 1: Range finding study results (cumulative mortality)

Nominal Loading Rate

(mg/L)

3 hours

6 hours

24 hours

48 hours

72 hours

96 hours

Control

0

0

0

0

0

0

1

0

0

0

0

0

0

10

0

0

0

0

0

0

100

0

0

0

0

0

0

 

Table 2: Definitive study results

Nominal Loading Rate

(mg/L)

Cumulative Mortality

% Mortality

3 hours

6 hours

24 hours

48 hours

72 hours

96 hours

96 hours

Control

0

0

0

0

0

0

0

100

0

0

0

0

0

0

0

 

Validity criteria fulfilled:
yes
Conclusions:
Under the conditions of the test, the 96 hour LL50 of the test material was determined to be greater than 100 mg/L. The NOELR was determined to be 100 mg/L.
Executive summary:

The acute toxicity to fish of the test material was determined in a study conducted in accordance with EU Method C.1 and OECD 203. Due to the low water solubility of the test material, the test solutions were prepared as a WAF. A range-finding study indicated no mortality up to a loading rate of 100 mg/L. The definitive test was conducted as a limit test with a loading rate of 100 mg/L.

Under the conditions of the test, the 96 hour LL50 of the test material was determined to be greater than 100 mg/L. The NOELR was determined to be 100 mg/L.

Description of key information

In the key study, the acute toxicity to fish of the test material was determined in a study conducted in accordance with EU Method C.1 and OECD 203. Due to the low water solubility of the test material, the test solutions were prepared as a WAF. A range-finding study indicated no mortality up to a loading rate of 100 mg/L. The definitive test was conducted as a limit test with a loading rate of 100 mg/L.

Under the conditions of the test, the 96 hour LL50 of the test material was determined to be greater than 100 mg/L. The NOELR was determined to be 100 mg/L.

Key value for chemical safety assessment

LC50 for freshwater fish:
100 mg/L

Additional information