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Short-term toxicity to aquatic invertebrates

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Reference
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
Experimental test result performed using standard test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
GLP compliance:
no
Analytical monitoring:
no
Vehicle:
yes
Remarks:
Acetone
Details on test solutions:
The stock solution 1.0 g/l was prepared in acetone. Test solutions of required concentrationas were prepared by mixing the stock solution of the test sample with reconstituted test water.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Water flea
- Strain: Straus
- Source: Own breeding at University of Chemistry and Technology, Prague
- Age at study initiation (mean and range, SD): The animals used for the test shall be less than 24 h old and should not be first brood progeny
- Feeding during test: No feeding

ACCLIMATION - No data available
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Remarks on exposure duration:
± 1 hr
Test temperature:
20±1°C
pH:
sample at concentration 2.0 mg.l'1: pH = 7.8 did not change during the test,
control: = 7.8 changed to pH = 7.9 during the test
control + acetone: 7.8 changed to pH = 7.9 during the test
Dissolved oxygen:
higher than 7.9 mg/L at the end of test
Nominal and measured concentrations:
Nominal test concentrations: 0.06 , 0.12 , 0.24 , 0.50 , 1.00 , 2.00 mg/l, respectively
Details on test conditions:
TEST SYSTEM
- Test vessel: 50 ml glass vessel
- fill volume: 25 ml
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water:
Natural water (surface or ground water), reconstituted water or dechlorinated tap water are acceptable as culturing and dilution water if D. magna survives in it for the duration of the culturing, acclimation and testing without showing signs of stress. Waters in the range pH 6 to pH 9, with hardness between 140 mg/l and 275 mg/l (as CaCO3) are recommended.
As an example, the preparation of dilution water meeting the requirements is described below.
Dissolve known quantities of reagents in water. The dilution water prepared shall have a pH of 7.8 ± 0.5, a hardness of (225 ± 50) mg/l (expressed as CaCO3), a molar Ca + Mg ratio close to 4 + 1 and a dissolved oxygen concentration above 7 mg/l.

Prepare the solutions specified below:
- Calcium chloride solution: Dissolve 117.6 g of calcium chloride dihydrate (CaCl2.2H2O) in water (4.2) and make up to 1 l with water (4.2).
- Magnesium sulfate solution: Dissolve 49.3 g of magnesium sulfate heptahydrate (MgSO4.7H2O) in water (4.2) and make up to 1 l with water (4.2).
- Sodium bicarbonate solution: Dissolve 25.9 g of sodium bicarbonate (NaHCO3) in water (4.2) and make up to 1 l with water (4.2).
- Potassium chloride solution: Dissolve 2.3 g of potassium chloride (KCI) in water (4.2) and make up to 1 l with water (4.2).

Mixing
Mix 2.5 ml of each of the four solutions and make up to 1 l with water.
The dilution water shall be aerated until the dissolved oxygen concentration has reached saturation and the pH has stabilized. If necessary, adjust the pH to 7.8 ± 0.5 by adding sodium hydroxide (NaOH) solution or hydrochloric acid (HCI). The dilution water prepared in this way shall not be further aerated before use.

- Sodium hydroxide solution, e.g. [NaOH] : 1 mol/l.
- Hydrochloric acid, e.g. [HCl] : 1 mol/l.

Reference substance:
Dissolve 600 mg of potassium dichromate (K2Cr2O7) in water and make up to 1 l with water (4.2).

OTHER TEST CONDITIONS
- Adjustment of pH: no adjustment done
- Photoperiod: No - Darkness
- Light intensity:

CALCULATION:
EC50 was calculated using non linear regression by the software Prism 4.0
Reference substance (positive control):
yes
Remarks:
Potassium dichromate (K2Cr2O7)
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
1.84 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95 % CI 1.06 - 3.17 mg/l
Results with reference substance (positive control):
- Results with reference substance valid
- EC50: 0.73 mg/L (24 hours)
Reported statistics and error estimates:
EC50 was calculated using non linear regression by the software Prism 4.0
Validity criteria fulfilled:
yes
Conclusions:
The median effective concentration (EC50) for the test substance on Daphnia magna was determined to be 1.84 mg/L on the basis of mobility inhibition effects in a 48 hour study.
Executive summary:

An acute immobilisation test was conducted for 48 hrs for assessing the short term toxicity of test chemical to aquatic invertebrate. Study was performed according to the OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test). Daphnia magna was used a test organism. The stock solution 1.0 g/l  was prepared in acetone. Test solutions of required concentrationas were prepared by mixing the stock solution of the test sample with reconstituted test water. 0.06 , 0.12 , 0.24 , 0.50 , 1.00 , 2.00 mg/l, respectively nominal concentrations were used in the study. Effects on immobilisation were observed for 48 hours and conducted under the static system. With the test substance one positive control Potassium dichromate (K2Cr2O7) was also run simultaneously. After the exposure of chemical, effect concentration EC50 was calculated using nonlinear regression by the software Prism 4.0. Based on the immobilisation of Daphnia magna due to the exposure of test chemical for 48 hours, the EC50 value was determine to be 1.84 mg/l. EC50 value indicates that the substance is likely to be hazardous to aquatic invertebrates and can be consider to be classified in aquatic chronic 2 category as per the CLP criteria.

Description of key information

An acute immobilisation test was conducted for 48 hrs for assessing the short term toxicity of test chemical to aquatic invertebrate. Study was performed according to the OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test). Daphnia magna was used a test organism. The stock solution 1.0 g/l  was prepared in acetone. Test solutions of required concentrationas were prepared by mixing the stock solution of the test sample with reconstituted test water. 0.06 , 0.12 , 0.24 , 0.50 , 1.00 , 2.00 mg/l, respectively nominal concentrations were used in the study. Effects on immobilisation were observed for 48 hours and conducted under the static system. With the test substance one positive control Potassium dichromate (K2Cr2O7) was also run simultaneously. After the exposure of chemical, effect concentration EC50 was calculated using nonlinear regression by the software Prism 4.0. Based on the immobilisation of Daphnia magna due to the exposure of test chemical for 48 hours, the EC50 value was determine to be 1.84 mg/l. EC50 value indicates that the substance is likely to be hazardous to aquatic invertebrates and can be consider to be classified in aquatic chronic 2 category as per the CLP criteria.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Effect concentration:
1.84 mg/L

Additional information

Based on the various experimental studies from different sources including secondary databases, authoritative database and studies were reviewed for the evaluation of the effects of test chemical and structurally and functionally similar read across chemicals on the immobility of aquatic invertebrates. The studies are as mentioned below:

An acute immobilisation test was conducted for 48 hrs for assessing the short term toxicity of test chemical to aquatic invertebrate. Study was performed according to the OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test). Daphnia magna was used a test organism. The stock solution 1.0 g/l  was prepared in acetone. Test solutions of required concentrationas were prepared by mixing the stock solution of the test sample with reconstituted test water. 0.06 , 0.12 , 0.24 , 0.50 , 1.00 , 2.00 mg/l, respectively nominal concentrations were used in the study. Effects on immobilisation were observed for 48 hours and conducted under the static system. With the test substance one positive control Potassium dichromate (K2Cr2O7) was also run simultaneously. After the exposure of chemical, effect concentration EC50 was calculated using nonlinear regression by the software Prism 4.0. Based on the immobilisation of Daphnia magna due to the exposure of test chemical for 48 hours, the EC50 value was determine to be 1.84 mg/l. EC50 value indicates that the substance is likely to be hazardous to aquatic invertebrates and can be consider to be classified in aquatic chronic 2 category as per the CLP criteria.

Above study further supported by the second study from authoritative database. Aim of this study was to assess the short term toxicity of test chemical to aquatic invertebrates. Study was performed according to the OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test). Daphnia magna was used as a test organism. Test chemical was analytically monitorized by HPLC method. 0, 1.0, 1.8, 3.2, 5.6, 10 mg/L respectively concentrations were used in the study. Test conducted under the static system on Juvenile (Less than 24 hours after birth) for 48 hours. Based on the effects observation on the mobility of Daphnia magna due to the test chemical exposure for 48 hours, the E50 value was determined to be 2.4 mg/l. This EC50 value indicates, that the test chemical was toxic to the aquatic invertebrate Daphnia magna and thus can be consider to be classified in aquatic chronic category 2 as per the CLP classification criteria.

Principle of this third weight of evidence study was to determine the short term toxicity of test chemical to aquatic invertebrates. Daphnia magna was used as a test organism. Syudy was performed under the static systme for 48 hours. (6 - 24) hours old Daphnia magna was used in the study which was collected from the laboratory. Based on the immobility of Daphnia magna, the EC50 value was observed to be 3.4 mg/l after exposure period of 48 hours, This EC50 value indicates that the test chemical was toxic to the aquatic invertebrate Daphnia magna and thus can be consider to be classified in aquatic chronic category 2 as per the CLP classification criteria.

Thus based on the above all studies and effects observations, it was concluded that the test chemical was toxic and classified in aquatic chronic category 2 as per the CLP classification criteria.