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Short-term toxicity to aquatic invertebrates

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short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2007-08-03 to 2007-09-13
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study
according to guideline
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
GLP compliance:
Analytical monitoring:
Details on sampling:
- Sampling method: Water samples were taken from each test chamber prior to test initiation (pre-test), at test initiation and at test termination. Water samples (10 mL) were collected using a 10 mL volumetric pipette and placed in a 20-mL glass scintillation vial.
Details on test solutions:

- Method: The nominal test concentrations were selected on the basis of the reported solubility of the substance (34 μg/L). A primary stock was prepared by isotropic dilution of the 14C-labeled L3 with non-radiolabelled L3. A 7.0 μL aliquot of 14C-L3 (0.00583 g) was added to a 1.0 mL vial. A 200 μL aliquot (0.16344 g) of L3 was also added to the vial. Approximately 0.5 mL of dimethyl formamide (DMF) was added to the vial and the contents of the vial was transferred to a 500-mL volumetric flask. The flask was then brought to volume with DMF, and mixed well. Three 0.1 mL aliquots of the primary stock were counted by LSC. The specific activity of the primary stock was determined to be 6.743 μCi/mg.

Four additional stock solutions were prepared at concentrations of 170, 85, 43 and 21 mg/L by proportional dilution of the primary stock with DMF. DMF only was used as the solvent control. The stock solutions were delivered to the diluter mixing chambers (at a rate of 0.0125 mL/minute) where they were diluted with dilution water (at a rate of 125 mL/min) to achieve the desired test concentrations. The concentration of DMF in the solvent control and the treated media was 0.1 mL/L. Test vessels and mixing chambers remained clear and colourless throughout the test.
Test organisms (species):
Daphnia magna
Details on test organisms:

- Source, supplier, any pre-treatment, breeding method:  Dow Corning in-house Cultures. The cultures were initiated with organisms obtained from Aquatic Biosystems Inc., Fort Collins, Colorado.      

- Age at study initiation:  <24 hours old

_ Culture conditions: Daphnia were cultured in dilution water at 20+/-2° under cool white fluorescent lighting with a 16:8 h light:dark photoperiod and a light intensity of 100+/-50 foot candles. They were fed on a diet of Selenastrum capricornutum and YCT (yeast, cerophyl and trout chow). Daphnids were not fed during the test.
Test type:
Water media type:
Limit test:
Total exposure duration:
48 h
Hardness 132 mg/L as CaCO3, Alkalinity 45 mg/L as CaCO3
Test temperature:
19.7 to 20.2ºC
Dissolved oxygen:
≥8.0 mg/L (90% ASV)
Not applicable
Nominal and measured concentrations:
Nominal concentrations: Negative Control, Solvent Control, 2.1, 4.3, 8.5, 17 and 34 μg/L 

Mean measured concentrations:
The mean measured concentrations in the treated media corresponded to 57, 49, 55, 56 and 59% of the nominal values respectively.

The results are reported with reference to mean measured concentrations.
Details on test conditions:

- Exposure vessel type:  The test compartments were 300-mL glass beakers with 2 holes on opposite sides covered with nylon screen.  The test chambers were placed in 7.5-L polyethylene aquaria containing 3.4 L of test solution.  

- Flow-through rate: Each replicate received approximately 26 volume additions per day delivered by rotameters. The rate of addition was set relatively high to compensate for the relatively high volatility of the test substance (vapour pressure 4 mm at 25°C).

- Dilution water source:  Dechlorinated Municipal Water

- Dilution water chemistry (hardness, alkalinity, pH, TOC, TSS, salinity, Ca/Mg ration, Na/K ratio):  Hardness 132 mg/L as CaCO3, Alkalinity 45 mg/L as CaCO3, pH 7.3-7.6, Conductivity 384  mhos/cm and Total Residual Chlorine <10  g/L

- Lighting (quality, intensity, and periodicity):  Wavelength similar to natural sunlight, 55 to 62 foot-candles, 16 hours light:8 hours dark

- Water chemistry in test:  DO, pH and temperature measured in each test chamber daily

- Element (unit) basis (i.e., immobilization):  Mortality and immobilization

- Test design (number of replicates, individuals per replicate, concentrations):  Two replicates/treatment, 10 individuals/replicate

- Method of calculating mean measured concentrations:  Arithmetic mean.
Reference substance (positive control):
48 h
Dose descriptor:
Effect conc.:
> 20 µg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
48 h
Dose descriptor:
Effect conc.:
>= 20 µg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
Details on results:
- Immobilisation of control: 0
Reported statistics and error estimates:
No significant effects on mobility or mortality were recorded in the highest treatment. An EC50 value could therefore not be determined.

Table 1. Test results

 Nominal concentration (μg/L)  Mean measured concentration (μg/L) Mean percentage immobilisation after 24 hours   Mean percentage immobilisation after 48 hours 
 0 (Control)  <LoQ  0  0
 0 (Vehicle control)  <LoQ  0  0
 2.1  1.2  0  0
 4.3  2.1  0  0
 8.5  4.7  0  0
 17  9.5  0  0
 34  20  0  5

The geometric mean of the highest concentration tested has been calculated to be 20 μg/L by the current reviewer.

Validity criteria fulfilled:
A 48-hour EC50 value of >20 μg/L and NOEC of ≥20 μg/L have been determined for the effects of the substance on mobility of Daphnia magna based on geometric mean measured concentrations. The results were obtained under flow-through test conditions.

Description of key information

Short-term toxicity to invertebrates: 48 h EC50 >20 μg/L (measured, geom. mean; highest concentration tested) (OECD 202).

Key value for chemical safety assessment

Additional information

A 48-hour EC50 value of >20 μg/L and NOEC of ≥20 μg/L (highest concentration tested) have been determined for the effects of the substance on mobility of Daphnia magna based on geometric mean measured concentrations. In view of the use of flow-through test conditions it is likely that the test organisms were exposed primarily to the parent substance.