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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:
key study
Study period:
14 September 2017 to 06 October 2017
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)
Deviations:
no
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
- Sampling method: Samples for possible analysis were taken from all test concentrations and the control at t=0 h and t=48 h. 4.8 mL was taken from the approximate centre of the test vessels.
- Sample storage conditions before analysis: Samples were stored at room temperature until analysis at the analytical laboratory of the Test Facility.
- At the end of the exposure period, the replicates were pooled at each concentration before sampling.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Preparation of test solutions started with a loading rate of 100 mg/L applying a 10-minute period of ultrasonic waves followed by a 3-day period of magnetic stirring to ensure maximum dissolution of the test material in medium. Thereafter, the aqueous Saturated Solution (SS) was collected by means of filtration through a 0.45 µm membrane filter (RC55, Whatman) and used as the highest test concentration. Lower test concentrations were prepared by subsequent dilutions of the SS in test medium. All test solutions were clear and colorless at the end of the preparation procedure.
- Any residual volumes were discarded.
- No correction was made for the purity/composition of the test material.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Strain/clone: Daphnia magna (Crustacea, Cladocera) (Straus, 1820), at least third generation, obtained by a cyclical parthenogenesis under specified breeding conditions.
- Source: In-house laboratory culture
- Daphnids originated from a healthy stock, 2nd to 5th brood, showing no signs of stress such as mortality >20 %, presence of males, ephippia or discoloured animals and there was no delay in the production of the first brood.
- Age: Young daphnids with an age of < 24 hours, from parental daphnids of more than two weeks old were selected for the test.
-Breeding was performed with newborn daphnids, i.e. less than 3 days old, by placing about 250 of them into 5 litres of medium in an all-glass culture vessel.
- Feeding during test: no
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
180 mg/L expressed as CaCO3
Test temperature:
19-20 °C
pH:
7.8-8.0
Dissolved oxygen:
9.3-9.6 mg/L
Nominal and measured concentrations:
Nominal: 1, 10 and 100 %
Measured: 100 % solution was 21 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: glass 100 mL vessel
- Type (delete if not applicable): open / closed
- Material, size, headspace, fill volume: 80 mL
- Aeration: no
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4 for the high concentration (100%), 2 for 1 and 10 %
- No. of vessels per control (replicates): 4
- Daphnids were introduced within 24 minutes after preparation of the test solutions.

TEST MEDIUM / WATER PARAMETERS
- Composition of medium M7:
Adjusted ISO medium: the following chemicals (analytical grade) are dissolved in tap water purified by Reverse Osmosis: Macro salts: CaCl2.2H2O 211.5 mg/L, MgSO4.7H2O 88.8 mg/L, NaHCO3 46.7 mg/L and KCl 4.2 mg/L.
Medium M7: trace elements, macronutrients and vitamins are added to freshly prepared ISO medium to reach the following concentrations: Trace elements: B 0.125 mg/L, Fe 0.05 mg/L, Mn 0.025 mg/L, Li, Rb and Sr 0.0125 mg/L, Mo 0.0063 mg/L, Br 0.0025 mg/L, Cu 0.0016 mg/L, Zn 0.0063 mg/L, Co and I 0.0025 mg/L, Se 0.0010 mg/L, V 0.0003 mg/L and Na2EDTA.2H2O 2.5 mg/L. Macro nutrients: Na2SiO3.9H2O 10.0 mg/L, NaNO3 0.27 mg/L, KH2PO4 0.14 mg/L and K2HPO4 0.18 mg/L. Vitamins: Thiamine 75.0 µg/L, B12 1.0 µg/L and Biotin 0.75 µg/L.
- Intervals of water quality measurement: at the beginning and the end of the test

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 hours photoperiod daily

EFFECT PARAMETERS MEASURED:
- Immobility (including mortality): At 24 hours and at 48 hours.
- pH and Dissolved oxygen: At the beginning and at the end of the test, for all concentrations and the control.
- Temperature of medium: Continuously in a temperature control vessel, beginning at the start of the test.
Reference substance (positive control):
yes
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 21 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
- Immobility: No daphnids became immobilised throughout the duration of the test.
- Experimental Conditions: The results of measurement of pH and oxygen concentrations (mg/L) remained within the limits prescribed by the study plan (pH: 6-9, not varying by more than 1.5 units; oxygen: ≥3 mg/L at the end of the test). The temperature continuously measured in a temperature control vessel varied between 19 and 20 °C during the test, and complied with the requirements as laid down in the study plan (18-22 °C, constant within 2 °C).
Results with reference substance (positive control):
- The reference test was carried out to check the sensitivity of the test system. Daphnia were exposed for a maximum of 48 hours to K2Cr2O7 concentrations of 0.10, 0.18, 0.32, 0.56, 1.0 and 1.8 mg/L and to a control. Twenty daphnids were exposed per concentration.
- In conclusion, the actual responses in this reference test with K2Cr2O7 are within the ranges of the expected responses at the different concentrations, i.e. the 48h-EC50 was between 0.3 and 1.0 mg/L. Hence, the sensitivity of this batch of D. magna was in agreement with the historical data collected at Charles River Den Bosch.
- The 24h-EC50 was 0.59 mg/L with a 95% confidence interval between 0.52 and 0.68 mg/L.
- The 48h-EC50 was 0.41 mg/L with a 95% confidence interval between 0.34 and 0.46 mg/L.
Reported statistics and error estimates:
No EC50 could be calculated because the test material proved to be non-toxic (EC50 > maximum soluble concentration tested).
Validity criteria fulfilled:
yes
Conclusions:
Under the conditions of this study, the 48h-EC50 for Daphnia magna exposed to the test material was beyond the range tested, i.e. exceeded a concentration of 21 mg/L, being considered the maximum soluble concentration of the test material in test medium.
Executive summary:

The potential of the test material to cause toxicity to aquatic invertebrates was determined in accordance with the standardised guideline OECD 202, under GLP conditions.

The objective of the study was to evaluate the test material for its ability to generate acute toxic effects on the mobility of Daphnia magna during an exposure period of 48 hours and, if possible, to determine the EC50 at 24 and 48 hours of exposure.

A Saturated Solution (SS) of the test material was prepared at a loading rate of 100 mg/L and used as the highest concentration. Lower concentrations were prepared by diluting the highest concentration in test medium.

A combined limit/range-finding test was performed. Twenty daphnids per group (5 per replicate, quadruplicate) were exposed to an untreated control and to the undiluted SS, in a limit test. In addition ten daphnids per group (5 per replicate, duplicate) were exposed to 1.0 and 10 % of the SS in the combined range-finding test. The total exposure period was 48 hours and samples for analytical confirmation of exposure concentrations were taken at the start and at the end of the test.

No daphnids became immobilised throughout the duration of the test.

Samples taken from the undiluted SS were analysed. The results were, however, not acceptable, since they were above the validated range. Consequently, the reserve samples were analysed. The actual exposure concentration was 21 mg/L at the start of the test. This concentration remained stable throughout the test duration, i.e. was 95 % of initial at the end of the test. Based on this results, effects parameters were based on the initially measured concentration.

Under the conditions of this study, the 48h-EC50 for Daphnia magna exposed to the test material was beyond the range tested, i.e. exceeded a concentration of 21 mg/L, being considered the maximum soluble concentration of the test material in test medium.

Description of key information

Under the conditions of this study, the 48h-EC50 for Daphnia magna exposed to the test material was beyond the range tested, i.e. exceeded a concentration of 21 mg/L, being considered the maximum soluble concentration of the test material in test medium.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Effect concentration:
21 mg/L

Additional information

The potential of the test material to cause toxicity to aquatic invertebrates was determined in accordance with the standardised guideline OECD 202, under GLP conditions. The study was awarded a reliability score of 1 in accordance with the criteria set forth by Klimisch et al. (1997).

The objective of the study was to evaluate the test material for its ability to generate acute toxic effects on the mobility of Daphnia magna during an exposure period of 48 hours and, if possible, to determine the EC50 at 24 and 48 hours of exposure.

A Saturated Solution (SS) of the test material was prepared at a loading rate of 100 mg/L and used as the highest concentration. Lower concentrations were prepared by diluting the highest concentration in test medium.

A combined limit/range-finding test was performed. Twenty daphnids per group (5 per replicate, quadruplicate) were exposed to an untreated control and to the undiluted SS, in a limit test. In addition ten daphnids per group (5 per replicate, duplicate) were exposed to 1.0 and 10 % of the SS in the combined range-finding test. The total exposure period was 48 hours and samples for analytical confirmation of exposure concentrations were taken at the start and at the end of the test.

No daphnids became immobilised throughout the duration of the test.

Samples taken from the undiluted SS were analysed. The results were, however, not acceptable, since they were above the validated range. Consequently, the reserve samples were analysed. The actual exposure concentration was 21 mg/L at the start of the test. This concentration remained stable throughout the test duration, i.e. was 95 % of initial at the end of the test. Based on this results, effects parameters were based on the initially measured concentration.

Under the conditions of this study, the 48h-EC50 for Daphnia magna exposed to the test material was beyond the range tested, i.e. exceeded a concentration of 21 mg/L, being considered the maximum soluble concentration of the test material in test medium.