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Diss Factsheets

Ecotoxicological information

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2015
Report date:
2015

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Deviations:
no
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Chemical structure
Reference substance name:
sodium 4-[(4-chlorobenzoyl)amino]benzoate
EC Number:
807-461-4
Cas Number:
1489170-67-3
Molecular formula:
C14H9ClNNaO3
IUPAC Name:
sodium 4-[(4-chlorobenzoyl)amino]benzoate
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
- Storage condition of test material: room temperature in the dark
- Description: white powder

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
- Concentrations: 100% v/v saturated solution test preparations (nominal)
- Sample storage conditions before analysis: The samples were stored frozen prior to analysis
- Preparation of Calibration Standards:
The test item (nominal 100 mg) was dissolved in 0.1 % (v/v) formic acid in methanol (100 mL) to prepare a stock solution with a nominal concentration of 1000 mg/L. This stock solution was further diluted with 0.1 % (v/v) formic acid in methanol to obtain a nominal 10 mg/L calibration standard. A duplicate calibration standard was similarly prepared at 10 mg/L. These duplicate calibration standards were used to determine the recovery and test sample concentrations.
- Preparation of Linearity Standards:
The test item (nominal 100 mg) was dissolved in 0.1 % (v/v) formic acid in methanol (100 mL) to prepare a stock solution with a nominal concentration of 1000 mg/L. Defined volumes of this stock solution were diluted with 0.1 % (v/v) formic acid in methanol to obtain standards in the range of 0.50 to 150 mg/L. A second standard was similarly prepared at a nominal concentration of 10 mg/L. These standards were used to evaluate the linearity of the analytical system.
- Preparation of Spiked Recovery Samples:
To demonstrate the validity of the analytical procedure, volumes of test medium were spiked with the test item and the recovery was assessed. The test item (nominal 100 mg) was initially dissolved in 0.1 % (v/v) formic acid in methanol to prepare a stock solution with a concentration of 1000 mg/L. A defined volume of this stock solution was diluted with test medium to obtain spiked recovery samples at a concentration of 1.0 mg/L. Five replicates were prepared and subjected to the same treatment as the test samples. In addition, test medium without the addition of the test item (synthetic control) was also analysed.
- Preparation of Test Samples:
The test samples were thawed with the aid of sonication. The test item was extracted from the test samples using a solid phase extraction cartridge (strata C18-E, 500 mg/ 3 mL). The cartridge was pre-conditioned with 10 mL of 0.1 % (v/v) formic acid in methanol and 10 mL of water. The samples were drawn through the cartridge under reduced pressure. Subsequently, the cartridge was eluted with 20 mL of 0.1 % (v/v) formic acid in methanol into a 20 mL volumetric flask. The solution was made up to the mark with 0.1 % (v/v) formic acid in methanol.

Test solutions

Vehicle:
yes
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Preliminary solubility work conducted indicated that it was not possible to obtain a testable solution of the test item using traditional methods of preparation e.g. ultrasonication and high shear mixing.
A preliminary media preparation trial indicated that a dissolved test item concentration of approximately 11 mg/L was obtained from a saturated solution method of preparation indicating this to be the limit of water solubility of this item under test conditions.
The test item solutions were prepared by stirring an excess (100 mg/L) of test item in test water using a propeller stirrer at approximately 1500 rpm for 24 hours. After the stirring period any undissolved test item was removed by filtration (0.2 µm Sartorius Sartopore filter, first approximate 1 L discarded in order to pre-condition the filter) to produce a 100% v/v saturated solution of the test item. A series of dilutions was made from this saturated solution to give further test concentrations of 10, 1.0 and 0.10% v/v saturated solution. Each prepared concentration was inverted several times to ensure adequate mixing and homogeneity.
Based on the results of the range-finding test the following test concentrations were assigned to the definitive test: 10, 18, 32, 56 and 100% v/v saturated solution.
- Controls: The control group was maintained under identical conditions but not exposed to the test item.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): Reconstituted water (ISO medium) used for both the range-finding and definitive test

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
The test was carried out using 1st instar Daphnia magna derived from in-house laboratory cultures.
Adult daphnia were maintained in 150 mL glass beakers containing Elendt M7 medium in a temperature controlled room at approximately 20 °C. The lighting cycle was controlled to give a 16 hours light and 8 hours darkness cycle with 20 minute dawn and dusk transition periods. Each culture was fed daily with a mixture of algal suspension (Desmodesmus subspicatus) and Tetramin® flake food suspension. Culture conditions ensured that reproduction was by parthenogenesis. Gravid adults were isolated the day before initiation of the test, such that the young daphnids produced overnight were less than 24 hours old. These young were removed from the cultures and used for testing. The diet and diluent water are considered not to contain any contaminant that would affect the integrity or outcome of the study.

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h

Test conditions

Hardness:
250 mg/L as CaCO3
Test temperature:
20-22 °C
pH:
7.8±0.2 (adjusted with NaOH or HCl)
Dissolved oxygen:
approximately air-saturation value
Nominal and measured concentrations:
Nominal concentrations of 10, 18, 32, 56 and 100% v/v saturated solution
Details on test conditions:
see "Any other information on materials and methods incl. tables"
Reference substance (positive control):
yes
Remarks:
potassium dichromate

Results and discussion

Effect concentrationsopen allclose all
Duration:
24 h
Dose descriptor:
EC50
Effect conc.:
> 10 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 10 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
24 h
Dose descriptor:
NOEC
Effect conc.:
10 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
10 mg/L
Nominal / measured:
meas. (not specified)
Conc. based on:
test mat.
Basis for effect:
mobility
Results with reference substance (positive control):
A positive control used potassium dichromate as the reference item at concentrations of 0.32, 0.56, 1.0, 1.8 and 3.2 mg/L.
Exposure conditions for the positive control were similar to those in the definitive test.
Analysis of the immobilisation data by the probit analysis using the linear maximum likelihood regression method at 24 and 48 hours using the ToxRat Professional computer software package based on the nominal test concentrations gave the following results:
EC50 (24h) = 1.1 mg/L, NOEC (24h) = 0.56 mg/L, LOEC (24h) = 1.0 mg/L
EC50 (48h) = 0.75 mg/L, NOEC (48h) = 0.56 mg/L, LOEC (48h) = 1.0 mg/L
The No Observed Effect Concentration is based upon equal to or less than 10% immobilisation at this concentration.
The results from the positive control with potassium dichromate were within the normal range for this reference item.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
A reliability of Klimisch 1 has been assigned as the acute toxicity of the test substance to aquatic invertebrates has been determined in a GLP test with Daphnia Magna according to OECD Guideline 202 and EU method C.2. Analysis of the 100% v/v saturated solution test preparations at 0 and 48 hours showed a measured test concentration of 10 mg/L was obtained.
Exposure of Daphnia magna to the test item gave EC50 values based on the measured test concentrations of greater than 10 mg/L after 24 and 48 h. The No Observed Effect Concentration (NOEC) was 10 mg/L after 24 and 48 h.
The substance does not need to be classified as Aquatic Acute Cat. 1, as the EC50 (48h) is > 1 mg/L. Further, although the EC50 (48h) is >1 and ≤10 mg/L, the substance does not need to be classified as Aquatic Chronic Cat. 2, as it is rapidly degradable and the log Pow is < 4 (criteria for substances for which adequate chronic toxicity data are not available).
The test was considered to be valid given that no more than 10% of the control daphnids showed immobilisation or other signs of disease or stress and that the oxygen concentration at the end of the test was >3 mg/L in the control and test vessels.