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Ecotoxicological information

Long-term toxicity to fish

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Link to relevant study record(s)

Reference
Endpoint:
fish, juvenile growth test
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Study period:
From May 04, 1995 to June 01, 1995
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
KL2 due to RA
Justification for type of information:
Refer to the section 13 for details on the category justification.
Reason / purpose for cross-reference:
read-across source
Qualifier:
according to guideline
Guideline:
OECD Guideline 204 (Fish, Prolonged Toxicity Test: 14-day Study)
Deviations:
no
Qualifier:
according to guideline
Guideline:
OECD Guideline 215 (Fish, Juvenile Growth Test)
Deviations:
yes
Remarks:
No inclusion of replicates
GLP compliance:
yes (incl. QA statement)
Analytical monitoring:
yes
Details on sampling:
- Concentrations: 0.1, 0.32 and 1 mg/L
- Sampling method: Duplicate samples were taken from approximate centre of the aquaria without mixing the test media of the control and of the of nominal 0.10 and 0.32 mg/L test concentrations at Day 0, 7, 14, 20 and 28 of the exposure period; From the test concentration of nominal 1.0 mg/L duplicate samples were taken on Days 0, 7, 14, 20 and 25, since after 25 days all fish were dead at this concentration.
- Determination of stability of the test substance in the stock solutions under the test conditions: Samples were taken from both freshly prepared stock solutions on Day 22 and just before the renewal of these stock solutions on Day 26
- Storage: Samples were preserved with 8% (v/v) of a Formalin solution (37%) until analysis
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Stock solution of the test substance was homogenized by intense stirring using magnetic stirrers (nominal concentration = 1.6 g/L)
- Dilution: The stock solution was diluted with deionized water to prepare the lower concentrated stock solution with nominal 0.16 g/L
- Storage: Continuously under refrigeration (at 3 - 8 deg C) to reduce the biological degradation
- Frequency: Stock solution was prepared for each test concentration every 2 to 4 days
- Eluate: 3.2 - 32 mL stock solution/h (20 dosages per h, 160 - 1600 microliter per dosage) of the stock solutions were dosed into mixing vessels using HAMILTON Digital dispenser (HAMILTON, Germany)
- Evidence of undissolved material (e.g. precipitate, surface film, etc): No remarkable observations were made concerning the behaviour of the test substance in the test media
Test organisms (species):
Oncorhynchus mykiss (previous name: Salmo gairdneri)
Details on test organisms:
TEST ORGANISM
- Common name: Rainbow trout
- Source: Forellenhof Fredelsloh, D-37186 Moringen
- Feeding during test: Yes

ACCLIMATION
- Acclimation period: 13 d
- Acclimation conditions (same as test or not): Yes

FEEDING DURING TEST
- Type of food: Commercial trout diet (Alleinfutter fur Forellenbrut, Aminostart (extr.), Kraftfutterwerk Wesel GmbH, D-46483 Wesel)
- Amount: Daily 4% dry weight (related to the mean initial fish wet weight)
- Feeding frequency: Daily
- Health during acclimation (any mortality observed): All fish were healthy
Test type:
flow-through
Water media type:
freshwater
Limit test:
no
Total exposure duration:
28 d
Hardness:
2.00-2.15 mmol/L (= 200-215 mg/L as CaC03), mean value: 2.1 0 mmol/L
Test temperature:
Ranged from 14.6 to 16.1°C (mean 15.0 to 15.7°C)
pH:
Ranged from 7.3 to 7.8

Dissolved oxygen:
> 7.6 mg/L (i.e. > 60%)

Salinity:
No data
Nominal and measured concentrations:
- Nominal: 0.032, 0.10, 0.32, 1.0 and 3.2 mg/L
- Measured concentrations of samples from 0.1, 0.32 and 1 mg/L nominal concentration solutions were 0, 0.79, 0.26 and 0.83 mg/L, respectively
Details on test conditions:
TEST SYSTEM
- Test vessel: Glass-aquarium
- Type (delete if not applicable): Open
- Material, size, headspace, fill volume: Glass-aquarium (length x width x height: 0.54 x 0.29 x 0.395 m) with 48 L test medium for each test concentration
- Renewal rate of test solution (frequency/flow rate): Daily; 16 L/h
- No. of organisms per vessel: 10 fish into each aquarium in a random order
- No. of vessels per concentration (replicates): Single
- No. of vessels per vehicle control (replicates): Single

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Drinking water of the community D-64380 Rossdorf
- Alkalinity: 4.55-4.80 mmol/L, mean value: 4.7 mmol/L

OTHER TEST CONDITIONS
- Adjustment of pH: Yes, adjusted to a pH of approximately 5
- Photoperiod: A 16-hour light to 8-hour darkness photoperiod with a 30 minute transition period between light and darkness
- Light intensity: 10-25 Lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Parameters: Mortality, symptoms of intoxication, body length and weight
- Frequency: Mortality and symptoms of intoxication: Daily; Growth parameters (i.e., body length and weight): At the start (Day 0) and end of the test (Day 28)
VEHICLE CONTROL PERFORMED: Yes

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.1-3.2
- Justification for using less concentrations than requested by guideline: Not applicable
- Range finding study
- Test concentrations: No data
- Results used to determine the conditions for the definitive study: Yes
Reference substance (positive control):
no
Key result
Duration:
28 d
Dose descriptor:
NOEC
Effect conc.:
ca. 0.32 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: mortalities and observed symptoms of intoxication
Remarks on result:
other: equivalent to a mean measured concentration of 0.26 mg/L
Key result
Duration:
28 d
Dose descriptor:
LOEC
Effect conc.:
ca. 1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: mortalities and observed symptoms of intoxication
Remarks on result:
other: equivalent to a mean measured concentration of 0.83 mg/L
Details on results:
Mortalities and symptoms of intoxication :
- Up to and including the test concentration of nominal 0.32 mg/L, no significant number of fish died during the test period of 28 days.
- At the test concentration of nominal 1.0 mg/L, all fish showed clear symptoms of intoxication and all died until Day 25.
- At the highest test concentration of nominal 3.2 mg/L all fish were dead after one day.

Effects on growth ((Refer to the Table 2 and 3) of the word document under 'attached background material') :
- At the start of the test, the mean values of body length and weight of the fish, placed in each test concentration and the control, were not significantly different. The mean body length of the test fish in the control had increased during the test period of four weeks by a factor of approximately 1.3, the mean body wet weight by a factor of approximately 3.1.
- According to the results of Williams-Tests (one-sided, α = 0.05), the mean body length and mean body wet weight of the surviving fish were statistically not significantly reduced compared to the control up to and including the test concentration of nominal 0.32 mg/L
- At the higher test concentrations of nominal 1.0 and 3.2 mg/L, the growth rates could not be determined, since all fish died during the test period.
Reported statistics and error estimates:
Analysis of variance (ANOVA) followed by multivariate one-sided Williams-Test

Analytical results: (refer to the table in the attached word document under 'attached background material')

Test solutionsThe analytically determined concentrations of the test substance in the analysed test media varied in the range from 61% to 112% of the nominal values. The mean measured values, calculated over all measurements during the test period amounted to 79% of nominal at the concentration of nominal 0.10 mg/L, to 81% at the concentration of nominal 0.32 mg/L, and to 83 % at the concentration of nominal 1.0 mg/L. Therefore, the biological results were based on the nominal concentrations as well on the mean measured test substance concentrations.

 

Stock solutionIn the stock solution samples, the measured concentrations amounted to in the mean 95% of the nominal values. The test substance was sufficiently stable in the stock solutions under the test conditions during the longest stock solution renewal periods of 4 days.

 

Discussion on observed results:

- The death of one fish at the test concentrations of nominal 0.10 and 0.32 mg test substance/L on Day 4 was not determined as a significant toxic effect of the test substance, since no clear symptoms of intoxication were observed at the test fish at these test concentrations, and according to the test guideline the death of one fish is tolerated even in the control.

- The reduced food uptake observed from Day 7 to 12 at the test concentration of nominal 0.32 mg/L (some uneaten food was observed at the bottom of the aquarium) had no significant effect on the growth rate of the test fish and was therefore not assessed as a significant toxic effect.

Validity criteria fulfilled:
not specified
Conclusions:
Under the study conditions, the nominal 28 d NOEC and LOEC for mortality, symptoms of toxicity and growth were established at 0.32 and 1.0 mg/L (equivalent to measured concentrations of 0.26 and 0.83 mg/L), respectively.
Executive summary:

A study was carried out to determine the chronic toxicity of the read-across substance, C12-18 and C18-unsatd. DEA, to Rainbow trout (Oncorhynchus mykiss) according to OECD Guidelines 204 and 215, in compliance with GLP. Ten fish per group were exposed to the test substance (dissolved in water using intense stirring) at 0.032, 0.10, 0.32, 1.0 and 3.2 mg/L for 28 d under flow-through conditions. Mortality and symptoms of toxicity were observed daily and the growth parameters (i.e. body length and weight) were recorded on Days 0 and 28. For analytical dose verification (HPLC-UV), samples were taken in the control, 0.10 and 0.32 mg/L groups on Days 0, 7, 14, 20 and 28 of exposure. At 3.2 mg/L, all fish were dead after one day. At 1.0 mg/L, all fish showed clear symptoms of toxicity and died by Day 25. No significant effect on mortality or signs of intoxication were observed up to the concentration of 0.32 mg/L. According to the results of a Williams test, the mean body length and mean body wet weight of the surviving fish were not significantly reduced compared to controls at up to and including the dose of 0.32 mg/L. Growth rates at higher concentrations could not be determined due to mortality. Under the study conditions, the nominal 28 d NOEC and LOEC for mortality, symptoms of toxicity and growth were established at 0.32 and 1.0 mg/L (equivalent to measured concentrations of 0.26 and 0.83 mg/L), respectively (Memmert, 1995).

Description of key information

Key value for chemical safety assessment

Fresh water fish

Fresh water fish
Dose descriptor:
NOEC
Effect concentration:
0.32 mg/L

Additional information

A study was carried out to determine the chronic toxicity of the read-across substance, C12-18 and C18-unsatd. DEA, to Rainbow trout (Oncorhynchus mykiss) according to OECD Guidelines 204 and 215, in compliance with GLP. Ten fish per group were exposed to the test substance (dissolved in water using intense stirring) at 0.032, 0.10, 0.32, 1.0 and 3.2 mg/L for 28 d under flow-through conditions. Mortality and symptoms of toxicity were observed daily and the growth parameters (i.e. body length and weight) were recorded on Days 0 and 28. For analytical dose verification (HPLC-UV), samples were taken in the control, 0.10 and 0.32 mg/L groups on Days 0, 7, 14, 20 and 28 of exposure. At 3.2 mg/L, all fish were dead after one day. At 1.0 mg/L, all fish showed clear symptoms of toxicity and died by Day 25. No significant effect on mortality or signs of intoxication were observed up to the concentration of 0.32 mg/L. According to the results of a Williams test, the mean body length and mean body wet weight of the surviving fish were not significantly reduced compared to controls at up to and including the dose of 0.32 mg/L. Growth rates at higher concentrations could not be determined due to mortality. Under the study conditions, the nominal 28 d NOEC and LOEC for mortality, symptoms of toxicity and growth were established at 0.32 and 1.0 mg/L (equivalent to measured concentrations of 0.26 and 0.83 mg/L), respectively (Memmert, 1995).