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Long-term toxicity to aquatic invertebrates

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Reference
Endpoint:
long-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
04 January to 18 May 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 211 (Daphnia magna Reproduction Test)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.20 (Daphnia magna Reproduction Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: 021340
- Expiration date of the lot/batch: 01 jULY 2022
- Purity test date: 25th August 2003

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature in the dark
- Stability under test conditions: stable
- Solubility and stability of the test substance in the solvent/vehicle: poorly soluble, stable
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: not stated

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: none
- Preliminary purification step (if any):none
- Final dilution of a dissolved solid, stock liquid or gel: WAFs prepared in modified Elendt M7 medium

Analytical monitoring:
yes
Details on sampling:
The test preparations were renewed daily throughout the test. The concentration and stability of the test item in the test preparation was verified by chemical analysis on Days 0, 6, 13 and 20 (fresh media) and on Days 1, 7, 14 and 21 (old media)
Vehicle:
yes
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: WAF's prepared
- Eluate: Modiefied Elendt M7 medium
- Differential loading:
- Controls: vehicle only
- Chemical name of vehicle (organic solvent, emulsifier or dispersant):
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): 0.1 - 5.0 mg/L nominal
- Evidence of undissolved material (e.g. precipitate, surface film, etc.):
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Dphnia magna
- Source: in-house laboratory culture
- Age of parental stock (mean and range, SD): 1st instar
- Feeding during test : Fed daily
- Food type: Each daphnid received approximately 5 to 15 µL of an algal suspension (Desmodesmus subspicatus) and approximately 20 µL of Tetramin® flake food suspension daily
- Amount: Feeding was at a level of approximately 0.1 to 0.2 mg carbon/daphnid/day, dependent on the age and size of the animals
- Frequency: Daily


Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
21 d
Hardness:
The water hardness of the control and the highest surviving test concentration in the fresh and old media was measured once per week. Water hardness was 1.8 mmol/L (= 180 mg/L as CaCO3)
Test temperature:
18 to 22 °C. The temperature was also measured every hour in one replicate of the control using a Testo temperature logger.
pH:
Dissolved oxygen concentrations, pH and temperature were recorded before and after each test media renewal. The pH and dissolved oxygen concentration were measured using a Hach HQ30d Flexi handheld meter whilst the temperature was measured using a Hanna Instruments HI 93510 digital thermometer. Measurements were made on one replicate for each test concentration.
Dissolved oxygen:
Dissolved oxygen concentrations, pH and temperature were recorded before and after each test media renewal. The pH and dissolved oxygen concentration were measured using a Hach HQ30d Flexi handheld meter whilst the temperature was measured using a Hanna Instruments HI 93510 digital thermometer. Measurements were made on one replicate for each test concentration.
Salinity:
N/a
Conductivity:
N/a
Nominal and measured concentrations:
Water samples were taken from the control and each surviving test group (replicates pooled) for quantitative analysis. Samples of the fresh test preparations were taken on Days 0, 6, 13 and 20 and of the expired test preparations on Days 1, 7, 14 and 21. Samples were stored frozen prior to analysis. Duplicate samples were taken and stored frozen for further analysis if necessary.
Details on test conditions:
TEST SYSTEM
- Test vessel:150 mL glass vessal
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: fill volume 100 mL
- Aeration: The test vessels were not aerated. The diluent water only was aerated prior to use.
- Type of flow-through (e.g. peristaltic or proportional diluter): N/a
- Renewal rate of test solution (frequency/flow rate): The test preparations were renewed daily throughout the test.
- No. of organisms per vessel: 1
- No. of vessels per concentration (replicates):10
- No. of vessels per control (replicates):10
- No. of vessels per vehicle control (replicates):N/a
- Biomass loading rate: N/a

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Deionised/purified water
- Total organic carbon:0
- Particulate matter:0
- Metals:
- Pesticides:
- Chlorine:
- Alkalinity:0.9 mmol/L
- Ca/mg ratio: 2.4
- Conductivity:
- Culture medium different from test medium:
- Intervals of water quality measurement:

OTHER TEST CONDITIONS
- Adjustment of pH: If necessary the pH was adjusted with NaOH or HCl to maintain a pH of7.9 ±0.3
- Photoperiod:16 hours light (not exceeding 1500 Lux) and 8 hours darkness with 20 minute dawn and dusk transition periods for 21 days
- Light intensity: see above

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.2
- Justification for using less concentrations than requested by guideline:
- Range finding study
- Test concentrations: 2.2
- Results used to determine the conditions for the definitive study:
Key result
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
0.45 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
immobilisation
Key result
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
immobilisation
Key result
Duration:
21 d
Dose descriptor:
LOEC
Effect conc.:
1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
immobilisation
Key result
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
0.45 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
reproduction
Key result
Duration:
21 d
Dose descriptor:
LOEC
Effect conc.:
1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
reproduction
Key result
Duration:
21 d
Dose descriptor:
EC10
Effect conc.:
0.54 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
reproduction
Key result
Duration:
21 d
Dose descriptor:
EL50
Effect conc.:
1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
reproduction
Details on results:
- Behavioural abnormalities:
- Observations on body length and weight: The results showed that there were no statistically significant differences (P≥0.05) between the control and the 0.045, 0.10, 0.22 and 0.45 mg/L test groups in terms of length of the daphnids after 21 days exposure to the test item, however, significant differences (P<0.05) were determined between the control and 1.0 mg/L loading rate WAF groups.
- Other biological observations:Daily observations throughout the test indicated there was an effect on the color of the surviving daphnids in the 1.0 mg/L loading rate which on occasions were observed to be paler in color than the control daphnids.
- Mortality of control:
- Other adverse effects control:
- Abnormal responses:
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values:
- Effect concentrations exceeding solubility of substance in test medium:
Validity criteria fulfilled:
yes
Conclusions:
The LOEL was 1.0 mg/L as there were increased mortalities in this test group compared to the control, this test group produced significantly fewer live young per adult (P<0.05) than the control group and the body lengths of the parental generation daphnids in this test group were significantly different (P<0.05) when compared to the control after 21 days.
The NOEL was 0.45 mg/L as there were no significant mortalities (immobilization) observed in the parental generation (P1), there were no significant differences (P≥0.05) in terms of the number of live young produced per adult and the body lengths of the parental generation daphnids in this test group were not significantly different when compared to the control after 21 days.

Description of key information

Key value for chemical safety assessment: 0.54 ( EL10

Key value for chemical safety assessment

EC10, LC10 or NOEC for freshwater invertebrates:
0.54 mg/L

Additional information