Registration Dossier

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
Study period:
2017-11-28 - 2017-11-30 (experimental phase)
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018
Report Date:
2018

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Version / remarks:
OECD Guideline for Testing of Chemicals No. 202, adopted 13. Apr. 2004: ”Daphnia sp., Acute Immobilisation Test“
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Version / remarks:
Council Regulation (EC) No. 440/2008, Method C.2. “Daphnia sp. Acute Immobilisation Test”, adopted 30. May 2008
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
yellow powder
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: sponsor

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: The test item was stored in a closed vessel dark and dry at room temperature (20 ± 5°C).

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
- Concentrations: 0, 10, 22, 46, 100, 220 mg/L
- Sample storage conditions before analysis: no

Test solutions

Vehicle:
no
Remarks:
stock solution dilution water
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
A stock solution containing 222 mg/L test item in dilution water was prepared. The lower concentrations were prepared by dilution of this stock solution with dilution water.

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: water flea
- Strain/clone: Daphnia magna STRAUS, strain Berlin
- Source: Origin: Umweltbundesamt Berlin, In-house breeding since 27. September 2007

Animal Husbandry
Daphnia magna is bred in the laboratory throughout the year. The animals are kept for the use in toxicity tests. They multiply by parthenogenesis, thus being genetically identical. The husbandry is performed similar to the method described in the OECD guideline.
Vessels: preserving glasses, nominal volume 2 L
Medium: M4-Medium (recipe of ELENDT)
Food: unicellular green algae (Desmodesmus subspicatus)
Medium renewal: twice a week
Photo period: 16/8 hours, using neon tubes
Temperature: 20 ± 2 °C

Selection of Daphnia
23 hours before the start of the test, the adult animals were separated from the young. 45 minutes before test start, the adults were caught with the help of a glass tube, and the newborn Daphnia, aged between 0 and 22.25 hours, were sieved from the medium and immediately placed into a beaker containing dilution water. After the settling-in period, animals which showed no apparent damage were used for the test.
Switching from M4-medium (husbandry) to Dilution water (test) has been shown not to cause any detrimental effects for test Daphnia.

Study design

Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Post exposure observation period:
none

Test conditions

Test temperature:
19.6 - 20.3°C
pH:
7.8
Dissolved oxygen:
8.7 - 8.9 mg/L
Nominal and measured concentrations:
Nominal: 0, 10, 22, 46, 100, 220 mg/L
Measured:
t=0h: 0, 10.2, 21.8, 43.6, 97.7, 224.86 mg/L
t=48h: 0, 11.2, 24.1, 48.7, 110.7, 255.00 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: glass beakers, nominal volume 50 mL, tall shape
- Renewal rate of test solution (frequency/flow rate): none
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Dilution water (Daphnia medium) with the following specification is used:
Parameter Concentration in mg/L
CaCl2*2H2O 293.80
MgSO4*7H2O 123.30
NaHCO3 64.80
KCl 5.80
Resulting hardness in mmol/L: 2.502
Resulting hardness in mg CaCO3/L: 250
Deviations from the nominal weighted loads were less than 5%. Exact values are recorded in the raw data. After preparation, the dilution water was aerated and the pH was measured. The pH was 7.8.

OTHER TEST CONDITIONS
- Photoperiod: 16/8 hours, using neon tubes

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Immobility: after 24 and 48 h
Test item concentration, pH, O2: at 0 and 48h
Reference substance (positive control):
yes
Remarks:
Potassium dichromate K2Cr2O7 (CAS No. 7778-50-9) was used as positive control in a current reference study.

Results and discussion

Effect concentrationsopen allclose all
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 220 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: no immobilisation occurred in any of the animals
Duration:
24 h
Dose descriptor:
EC50
Effect conc.:
> 220 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: no immobilisation occurred in any of the animals
Key result
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
>= 220 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: no immobilisation occurred in any of the animals
Duration:
48 h
Dose descriptor:
LOEC
Effect conc.:
> 220 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: no immobilisation occurred in any of the animals
Details on results:
- Mortality of control: no immobility occurred
- Other adverse effects control: none stated
- Abnormal responses: none observed
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: none
- Effect concentrations exceeding solubility of substance in test medium: none
Results with reference substance (positive control):
- Results with reference substance valid? yes
- Relevant effect levels: 24h-EC50i = 1.6 mg/L
- Limit test: no
- Dose-response test: yes
- ECx: 24h-EC50i = 1.6 mg/L
- Other:
In a current reference study, the 24h-EC50i value was determined as 1.6 mg/L. The value was within the requested range for the 24h-EC50 between 0.6 - 2.1 mg/L of potassium dichromate K2Cr2O7 as specified in the OECD guideline. Since the fits showed sufficient statistical correspondence of the data with the dose-response-equation, this reference study can be considered valid.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
The study was conducted under GLP according to OECD guideline 202 and EU method C.2 on the registered substance itself. The method is to be considered scientifically reasonable with no deficiencies in documentation or deviations from the guidelines, the validity criteria were met. Hence, the results can be considered as reliable to assess the toxicity of ZPS towards daphnids.
The study was performed using 5 concentrations ranging from 10 to 220 mg/L. For each test concentration, 20 Daphnia were exposed to the test item for 48 hours in a static test system. After 24 and 48 hours, the immobilised Daphnia were counted.
None of the concentrations showed toxicity (only one Daphnia was immobilised after 48h at 10 mg/L) and also none of the animals was immobilised in the blank control.
Potassium dichromate K2Cr2O7 (CAS No. 7778-50-9) was used as positive control in a current reference study to assure that the test conditions are reliable.
At the beginning and at the end of the test, the content of the test item in the test solutions was determined using HPLC-UV determination. The concentrations determined at the start of the test were between 94.7 and 102.3% of the nominal concentration. At the end of the test the determined concentrations were between 105.9 and 115.9% of the nominal concentrations. Therefore, the determination of the biological results was based on the nominal concentration.
No observations were made which might cause doubts concerning the validity of the study outcome. All validity criteria were met.
The result of the test is considered valid.
The EC50 > 220 mg/L is above the limit value for classification as aquatic chronic Cat. 3, so ZPS does not need to be classified according to Regulation 1272/2008 and amendments.
Executive summary:

The study was conducted under GLP according to OECD guideline 202 and EU method C.2 on the registered substance itself.

One valid experiment was performed.

The study was performed using 5 concentrations ranging from 10 to 220 mg/L. For each test concentration, 20 Daphnia were exposed to the test item for 48 hours in a static test system. After 24 and 48 hours, the immobilised Daphnia were counted.

None of the concentrations showed toxicity (only one Daphnia was immobilised after 48h at 10 mg/L) and also none of the animals was immobilised in the blank control.

Potassium dichromate K2Cr2O7 (CAS No. 7778-50-9) was used as positive control in a current reference study to assure that the test conditions are reliable.

At the beginning and at the end of the test, the content of the test item in the test solutions was determined using HPLC-UV determination. The concentrations determined at the start of the test were between 94.7 and 102.3% of the nominal concentration. At the end of the test the determined concentrations were between 105.9 and 115.9% of the nominal concentrations. Therefore, the determination of the biological results was based on the nominal concentration.

The following results were determined for the test item 3-(2-Benzothiazolylthio)-propanesulfonic acid, sodium salt (ZPS) (species: Daphnia magna).

48h-NOEC ≥ 220 mg/L
48h-LOEC > 220 mg/L
24h-EC50 > 220 mg/L
48h-EC50 > 220 mg/L