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Short-term toxicity to fish

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Reference
Endpoint:
short-term toxicity to fish
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Sep 21, 2018 - Nov 02, 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 203 (Fish, Acute Toxicity Test)
Version / remarks:
1992
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method C.1 (Acute Toxicity for Fish)
Version / remarks:
Council Regulation (EC) No. 440/2008 laying down test methods pursuant to Regulation (EC) No. 1907/2006 of the European Parliament and the council on the Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH).
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Specific details on test material used for the study:
TREATMENT OF TEST MATERIAL PRIOR TO TESTING
The test medium (reconstituted water and test item) was prepared freshly. Therefore, the calibrated flask with test medium was treated in an ultrasonic device for 1 hour. Subsequently, the preparation was stirred with a magnetic stirrer for further 23 hours. Afterwards, the formulation was passed through a membrane filter with a pore size of 0.2 µm. The filtrate was used for the study.
Analytical monitoring:
no
Vehicle:
yes
Details on test solutions:
TREATMENT OF TEST MATERIAL PRIOR TO TESTING
The test medium (reconstituted water and test item) was prepared freshly. Therefore, the calibrated flask with test medium was treated in an ultrasonic device for 1 hour. Subsequently, the preparation was stirred with a magnetic stirrer for further 23 hours. Afterwards, the formulation was passed through a membrane filter with a pore size of 0.2 µm. The filtrate was used for the study.

VEHICLE:
For the reconstituted water the following chemicals (analytical grade) were dissolved in fully demineralized water:

Macro nutrients mg/L
CaCl2 x 2H2O 294.00
MgSO4 x 7H2O 123.25
NaHCO3 64.75
KCl 5.75

Before the start of the experimental part the reconstituted water for fish was prepared. After preparation the water was aerated for 24 hours before using it.
The proportion of Ca to Mg ions was 4:1 and that of Na to K ions 10:1. Hardness: about 228 mg/L CaCO3.
Test organisms (species):
Danio rerio (previous name: Brachydanio rerio)
Details on test organisms:
TEST ORGANISM
- Common name: Zebrafish
- Strain: Dania rerio
- Origin: Max-Planck-Institut, Tübingen, Germany
- Breeder: Institute of Toxicology, Merck KGaA
- Length at study initiation: 2.7 - 2.9 cm
- Weight at study initiation: 0.2 - 0.4 g
- Feeding during test: commercial fish diet during acclimatization

ACCLIMATION
- Acclimation period: The fish were acclimatized to the conditions of the laboratory for more than 14 days and were held in reconstituted water for at least 12 days before testing.
- Acclimation conditions (same as test or not): same
- Health during acclimation (any mortality observed): no
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
96 h
Post exposure observation period:
no
Hardness:
228 mg/L CaCO3
Test temperature:
20.5 to 21.5°C
pH:
7.49 to 8.00
Dissolved oxygen:
72.8 - 93.1%
Conductivity:
710 µS/cm
Nominal and measured concentrations:
Nominal Concentration: 100 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: 10L Glass Aquariums
- Type (delete if not applicable): open
- Material, size, headspace, fill volume: Full glas folled with 10L test medium
- Aeration: no
- Type of flow-through (e.g. peristaltic or proportional diluter): static
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: reconstituted watr according OECD 203
- Total organic carbon: no data
- Particulate matter: no data
- Metals: no data
- Pesticides:no data
- Chlorine: no data
- Alkalinity: no data
- Ca/mg ratio: 4/1
- Culture medium different from test medium: no
- Intervals of water quality measurement: daily

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 12hrs light / 12hrs dark
- Light intensity: no data

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): mortality, clinical symptoms

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 100 mg/L
- Results used to determine the conditions for the definitive study: mo mortality observed at 100 mg/L
Reference substance (positive control):
no
Key result
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality (fish)
Remarks on result:
other: > 0.0000001448 mg/L (limit of water solubility)
Details on results:
The 96 hour LC50 for the test material to Zebrafish could not be quantified due to the absence of toxicity at the highest test concentration achievable in water. An aqueous filtrate of 0.1 g/L nominally revealed no aquatic toxicity in the test system.


The 96h LC50 exceeded the water solubility of 0.0000001448 mg/L (nominal >100 mg/L) and, thus, could not be determined in this study.


- Behavioural abnormalities: no
- Observations on body length and weight: no
- Other biological observations: no
- Mortality of control: no
- Other adverse effects control: no
- Abnormal responses: no
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: no
- Effect concentrations exceeding solubility of substance in test medium: yes
Validity criteria fulfilled:
yes
Conclusions:
Under the conditions of the present study, an aqueous solution of 100 mg/L of the test item revealed no aquatic toxicity in the test system.The 96h LC50 is greater than the maximum achievable concentration of 0.0000001448 mg/L (nominal > 100 mg/L) and, thus, could not be determined in this study.
Executive summary:

The objective of this study was to determine the acute toxicity of the test material in the zebrafish (Daniorerio).

For this purpose, 10 fish (test item group) were exposed to a nominal test item concentration of 100 mg/L (limit test) over 96 hours, under defined conditions in an open static system. Additionally, one control group (10 fish) was used.The fish were observed for signs of toxicity or mortality for 96 hours.

The test item concentration in the reconstituted water was not quantified at the start and the end of this study due to the low water solubility (0.0000001448 mg/L calculated). Because of the low water solubility, the compound cannot be detected with standard analytical methods. The development of an analytical method with a sufficiently low detection and quantification limit is complex. Due to the absence of any adverse effects at the saturation concentration, the study was performed without analytical concentration verification.After exposure of zebrafish (Danio rerio) to a nominal concentration of 100 mg/L of the test item for 96 hours no signs of toxicity and no mortality were observed.

For the test material, the following LC50values for zebrafish were determined.

24 h LC50      > 0.0000001448 mg/L (nominal > 100 mg/L)

48 h LC50      > 0.0000001448 mg/L (nominal > 100 mg/L)

72 h LC50      > 0.0000001448 mg/L (nominal > 100 mg/L)

96 h LC50      > 0.0000001448 mg/L (nominal > 100 mg/L)

Under the conditions of the present study, an aqueous solution of 100 mg/L of the test item revealed no aquatic toxicity in the test system.The 96h LC50 is greater than the maximum achievable concentration of 0.0000001448 mg/L (nominal > 100 mg/L) and, thus, could not be determined in this study.

Description of key information

OECD 203: LC50 (96 h)  > 0.0000001448 mg/L (limit of water solubility), 96 -h LC50 > 100 mg/L (nominal)

Key value for chemical safety assessment

Additional information

No adverse effects on fish were observed up to the limit of water solubility (0.0000001448 mg/L). Therefore, the 96 -h LC50 is greater than 100 mg/L (nominal concentration).