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Ecotoxicological information

Short-term toxicity to aquatic invertebrates

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Link to relevant study record(s)

Reference
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2016-06-07 to 2016-06-09
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Version / remarks:
13 April 2004
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Version / remarks:
30 May 2008
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 850.1010 (Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids)
Version / remarks:
April 1996
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Remarks:
TOXI-COOP Toxicology Research Center Zrt.
Analytical monitoring:
no
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The test concentrations were obtained by serial dilution of a saturated stock solution (100 mg/L loading rate). A supersaturated solution (nominal loading: 100 mg/L) was prepared by adding an excess of test item in ISO medium. An amount of 0.03 g test item was diluted in 300 mL ISO medium. Subsequently, this solution was shaken for 24 hours and thereafter the non-dissolved test material was removed by filtration through a 0.22 μm membrane filter to obtain the saturated stock solution (i.e. 100 % v/v saturated solution). The test solutions of chosen test concentrations were prepared by appropriate dilution of this stock solution. The test solutions were prepared immediately before introduction of daphnids.
- Controls:
Untreated Control: The dilution water (ISO-medium) was used without addition of the test item.
Toxic Reference Control: For the evaluation of the quality of the Daphnia clone and the experimental conditions, Potassium dichromate is tested at least twice a year to demonstrate satisfactory test conditions.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Water flea
- Strain/clone: Straus
- Source: Laboratory of Hydrobiology (Central Agricultural Office, Directorate of Plant-, and Soil Protection) 2100 Gödöllő, Kotlán S. u. 3. Hungary
- Age of parental stock: Less than 24 hours old
- Feeding during test: None

ACCLIMATION
- Acclimation period and conditions: Daphnia were bred in the Laboratory of TOXI-COOP ZRT. under similar temperature and light conditions as in the test, and in reconstituted water of a similar quality regarding to pH, components of the main ions and total hardness as the test water used in the test. Therefore, additional acclimatisation was not necessary.
- Type of food: The test animals were fed with centrifuged green alga suspension.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
249 mg/L (as CaCO3)
Test temperature:
19.2 – 19.7 °C
pH:
7.35 – 8.05
Dissolved oxygen:
7.79 – 8.07 mg/L
Nominal and measured concentrations:
Nominal: 1.25, 2.5, 5, 10 and 20 mg/L
No measured concentrations.
Details on test conditions:
TEST SYSTEM
- Test vessel: Glass beakers of approximately 40 mL test medium
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- ISO medium according to OECD 202 was used as dilution water in the experiment

OTHER TEST CONDITIONS
- Adjustment of pH: No, pH of the test solution was not adjusted and not varied by more than 1.5 units.
- Photoperiod:16 hours light (artificial illumination) and 8 hours darkness.

EFFECT PARAMETERS MEASURED: The immobility or mortality of the Daphnia was determined by visual observation after 24 and 48 hours of exposure. Those animals not able to swim within 15 seconds after gentle agitation of the test beaker were considered to be immobile.

RANGE-FINDING STUDY
- Test concentrations: 0, 0.1, 1, 10 and 100 mg/L
- Results used to determine the conditions for the definitive study:
Immobilisation Test:
After 48 hours no immobilised animals were determined in the control group and at 0.1 and 1 mg/L. At 10 mg/L 8 out of 10 animals were immobilised and at the highest test concentration of 100 mg/L all animals were immobile.

Preliminary analytical investigations:
Based on the measured data obtained during the preliminary analytical investigations the test item undergoes a fast hydrolysis reaction in the presence of 20% water. Its concentration decreased below 10% of the nominal within 13 minutes. Stability of the hydrolysis product, 1,4-Phenylenediamine was investigated under the same conditions. This compound also decomposed rapidly, less than 20% of the nominal concentration was measured within 13 minutes.
Consequently it was not possible to validate a suitable analytical method for the verification of test item concentration in aqueous test media. Analytical determination of the test item concentration was not feasible due to instability of the test item. Therefore, biological endpoints of the test are based on the nominal test item concentrations (loading rate).
Reference substance (positive control):
yes
Remarks:
Potassium dichromate (K2Cr2O7)
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
9.1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95% CL: 7.7 - 10.6
Duration:
48 h
Dose descriptor:
EC10
Effect conc.:
6.7 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95% CL: 4.2 - 7.9
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
5 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
LOEC
Effect conc.:
10 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Details on results:
- Behavioural abnormalities: No immobilisation or any abnormal behaviour of test animals was observed in the concentrations of 1.25, 2.5 and 5 mg/L after 48 hours of exposure.
- Mortality of control: None
- Other adverse effects control: None
Results with reference substance (positive control):
- Results with reference substance valid? Yes
- EC50 (24h): 1.26 mg/L (95% conf. limits: 1.11 – 1.55 mg/L)
Reported statistics and error estimates:
The 24-h and 48-h ECx values were calculated by Probit analysis using SPSS PC+ software.
The NOEC, LOEC values were determined directly from the raw data.
All biological results are related to the nominal test item concentrations.
Validity criteria fulfilled:
yes
Conclusions:
In this 48-hour acute toxicity test on Daphnia magna an EC50 of 9.1 mg/L was observed.
Executive summary:

An acute Daphnia magna Immobilisation Test was performed according to OECD Guideline 202. The purpose of this study was to evaluate the influence of the test item on the mobility respectively survival of Daphnia magna.

Young Daphnia were exposed to test item concentrations of 1.25, 2.5, 5, 10 and 20 mg/L (nominal) for 48 hours under static conditions. The test concentrations were obtained by serial dilution of a saturated stock solution (100 mg/L loading rate). Due to the rapid hydrolysis of the test item and its hydrolysis product (1,4-Phenylenediamine), validation of suitable analytical method was not possible for the verification of test item concentration in aqueous test media. Analytical determination of the test item concentrations was not feasible, therefore biological endpoints of the test are based on the nominal test item concentrations (loading rate). No immobilisation or any abnormal behaviour of test animals was observed in the control and in the concentrations of 1.25, 2.5 and 5 mg/L after 48 hours of exposure. Significant immobilisation was observed in the two highest test concentrations of 10 and 20 mg/L (65 and 100 % immobilisation respectively) at the end of the test.The determined 48h-EC50 was 9.1 (95% CL: 7.7. - 10.6).

Description of key information

In an 48-hour acute toxicity test on Daphnia magna an EC50 of 9.1 mg/L was observed.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Effect concentration:
9.1 mg/L

Additional information

An acute Daphnia magna Immobilisation Test was performed according to OECD Guideline 202. The purpose of this study was to evaluate the influence of the test item on the mobility respectively survival of Daphnia magna.

Young Daphnia were exposed to test item concentrations of 1.25, 2.5, 5, 10 and 20 mg/L (nominal) for 48 hours under static conditions. The test concentrations were obtained by serial dilution of a saturated stock solution (100 mg/L loading rate). Due to the rapid hydrolysis of the test item and its hydrolysis product (1,4-Phenylenediamine), validation of suitable analytical method was not possible for the verification of test item concentration in aqueous test media. Analytical determination of the test item concentrations was not feasible, therefore biological endpoints of the test are based on the nominal test item concentrations (loading rate). No immobilisation or any abnormal behaviour of test animals was observed in the control and in the concentrations of 1.25, 2.5 and 5 mg/L after 48 hours of exposure. Significant immobilisation was observed in the two highest test concentrations of 10 and 20 mg/L (65 and 100 % immobilisation respectively) at the end of the test.The determined 48h-EC50 was 9.1 (95% CL: 7.7. - 10.6).