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Short-term toxicity to aquatic invertebrates

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Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
6 Dec 2012 - 2 Jan 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
yes
Details on sampling:
- Concentrations: concentrations of dissolved cerium were determined in at least one of the triplicate samples from each treatment per sampling time
- Sampling method: triplicate samples were taken from each treatment at the start and end of each test medium renewal period (after filtration through a membrane filter, Whatman, Type NC45, pore size 0.45 µm).
- Sample storage conditions before analysis: at ambient temperature in the dark until used for testing in the analytical phase
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Due to the low solubility of the test item in test water, a dispersion of the test item with the loading rate of 101 mg/L (corresponding to 100 mg/L cerium ammonium nitrate corrected for purity) was sonicated for 15 minutes. After adjusting the pH to 6.0 using 1 M sodium hydroxide solution, the dispersion was continuously stirred at room temperature in the dark over 96 hours and then filtered through a membrane filter (Whatman, Type NC45, pore size 0.45µm).No auxiliary solvent or emulsifier was used.
The undiluted filtrate was used as highest concentrated test medium and as a stock solution for preparation of the test media with lower test concentrations.

Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia magna
- Strain: Straus
- Source: University of Sheffield / UK in 1992
- Age at study initiation (mean and range, SD): 6-24 hours, not first brood progeny
- Feeding during test: no
- Method of breeding: clone has been bred at Harlan Laboratories in reconstituted water of the quality identical to the water quality used in the test

ACCLIMATION
- Type and amount of food: green algae Desmodesmus subspicatus CHODAT, Strain No. 86.81 SAG, supplied by the Collection of Algal Cultures or a mixture of this algal suspension and a commercial fish diet
- Feeding frequency: three times a week
- Health during acclimation (any mortality observed): good. No mortalities observed
Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
250 mg CaCO3/L
Test temperature:
20°C-21°C
pH:
New test media:
0 h = 6.0-6.1
24 h = 6.0-6.1

Old test media:
24 h = 6.2-6.4
48 h = 6.7-7.2
Dissolved oxygen:
8.2 mg/L - 8.7 mg/L
Salinity:
not applicable
Nominal and measured concentrations:
Analytically determined concentrations in the test media (control, dilutions 1:16, 1:8, 1:4, 1:2, and the undiluted filtrate) were as follows:
Day 0, 0 h (beginning of experiment):
< LOQ, Day 1, 24 h (end of 1st renewal period):
< LOQ, < LOQ, < LOQ, < LOQ, 0.00108 and 0.00259 mg dissolved Ce/L corresponding to Day 1, 0 h (beginning of 2nd renewal period):
< LOQ, < LOQ, < LOQ, < LOQ, 0.00165 and 0.00336 mg dissolved Ce/L corresponding to Day 2, 24 h (end of 2nd renewal period):
< LOQ, < LOQ, < LOQ, < LOQ, 0.00167 and 0.00319 mg dissolved Ce/L corresponding to
At the start and end of the test, the analytically determined concentrations of dissolved cerium in the test media (dilutions 1:16, 1:8, 1:4, 1:2 and the undiluted filtrate) were below 4 μg/L, corresponding to concentrations of cerium ammonium nitrate below 14 μg/L.

To get an indication of the average concentration of dissolved cerium / cerium ammonium nitrate the daphnids were exposed to in this test, the mean measured concentrations were calculated for both dissolved cerium and cerium ammonium nitrate. These means were calculated in two steps. First, for each renewal period, the geometric mean was calculated from the concentrations measured at the beginning and end of the renewal period. Second, the arithmetic mean was
calculated from the geometric means of the two renewal periods.

The mean measured dissolved cerium concentratin (µg/L) in the test media (dilutions 1:16, 1:8, 1:4, 1:2) were as follows:
Details on test conditions:
TEST SYSTEM
- Test vessel: 100-mL glass beakers filled with 50 mL of test medium
- Type (delete if not applicable): covered with glass plates to reduce the loss of water by evaporation and to avoid the entry of dust into the solutions
- Aeration: aeration prior to the start of the study, no aeration during the test period
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- Biomass loading rate: 2mL test medium per daphnia

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Reconstituted test water according to ISO 6341
- Ca/mg ratio: 4:1
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Adjustment of pH: yes. Adjustment of dispersion to pH 6.0 after the 3-h stirring period using 1 M NaOH, 2) adjustment of test medium (for dilution of the filtrate) to pH 6.0 using 1 M HCl
- Photoperiod: A 16-hour light to 8-hour dark cycle with a 30-minute transition period was used.
- Light intensity: between 390 and 560 Lux.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : immobilisation, daily

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.
- Range finding study: yes
- Test concentrations: undiluted filtrate, 1:10, 1:100, 1:1000 dilution, and control
- Results used to determine the conditions for the definitive study: 0% immobilisation at 1:10 dilution, 100% immobilisation in undiluted filtrate
Reference substance (positive control):
yes
Remarks:
potassium dichromate
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 26 mg/L
Nominal / measured:
nominal
Conc. based on:
element
Remarks:
Ce
Basis for effect:
mobility
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
other: anhydrous cerium ammonium nitrate
Basis for effect:
mobility
Details on results:
After 24 and 48 hours of exposure, no immobilized test organisms were recorded in any of the treatments. The effect concentrations are therefore expressed based on nominal concentrations. The 24- and 48-hour EC0 and NOEC were both determined to be ≥ 26 and ≥ 100 mg/L for
dissolved cerium and cerium ammonium nitrate, respectively.
Results with reference substance (positive control):
- Results with reference substance valid? yes
- EC50/LC50: 48-hour EC50: 0.94 mg/L, was within historical range (from 2000 to 2012: 0.43-1.1 mg/L)
Reported statistics and error estimates:
NOEC, EC0 and EC100 were determined directly from the raw data.
24-h EC50 could not be calculated due to absence of toxicity and was therefore also determined directly from the raw data.
48-h EC50 and 95% confidence limits were calculated using Weibull Analysis with linear weighted regression.
The two lowest treatments were excluded from the statistical evaluation, as they did not contribute to the dose-response relationship.
Validity criteria fulfilled:
yes
Conclusions:
Cerium ammonium nitrate had no acute toxic effects on Daphnia magna under the conditions of the test up to the solubility limit of the test item in the test water (loading rate of 101 mg/L). The 48 hour NOEC and EC50 were therefore >= 100 mg/L and > 100 mg/L cerium ammonium nitrate respectively.
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
20 Dec 1994 - 22 Dec 1994
Reliability:
3 (not reliable)
Rationale for reliability incl. deficiencies:
other: The study is GLP-compliant and performed according to an internationally recognised guideline.
Remarks:
The results can however not be considered reliable. Dissolved Ce analysis indicates that Ce precipitates and disappears from the test solution. However, effect concentrations were based on nominal concentrations.
Qualifier:
according to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
- Test solution samples were analysed at 0 and 24 hours (separate bottles without test organisms kept under similar conditions).
Vehicle:
no
Details on test solutions:
- test solutions were prepared by adding test substance to the medium to obtain a final concentration of 100 mg/L
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: waterflea
- Strain: Strauss 1820 (IRCHA)
- Source: laboratory breed, laboratory of Ecotoxicology of CRIT/D
- Age at study initiation (mean and range, SD): not reported
- Size at study initiation (length definition, mean, range and SD): between 560 µm and 800 µm
- Feeding during test: no
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
48 h
Hardness:
Not reported.
Test temperature:
19.5-20 °C
pH:
8.1-8.5
Dissolved oxygen:
94-97 %
Salinity:
Not applicable.
Nominal and measured concentrations:
Nominal concentration: 100 mg/L
Measured concentration (in test solutions without test animals, kept under similar conditions): 104.2 mg test substance/L at t=0, 24 mg Ce/L at t=0, <= 3 mg Ce/L after 24 h. Test substance was clearly present in the precipitate at the bottom of the flasks.
Details on test conditions:
TEST SYSTEM
- Test vessel: 100-mL Pyrex beakers, 40 mL fill-volume
- No. of organisms per vessel: 20
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 1

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: bidistilled water
- Composition of test medium:
* 0.297 g/l CaCl2.2H2O
* 0.167 g/l MgCl2.6H2O
* 0.200 g/l NaHCO3
* 0.026 g/l K2SO4
- Conductivity: 0.90 µS/cm (dilution water)
- Ca:Mg ratio: 2.5:1

OTHER TEST CONDITIONS
- Adjustment of pH: not reported
- Photoperiod: continuous darkness

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): mobility, daily

TEST CONCENTRATIONS
- Spacing factor for test concentrations: limit test
- Range finding study: no
Reference substance (positive control):
yes
Remarks:
(potassium dichromate)
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
- EC50(24h) > 100 mg/L
- After 24 hours a decrease of dissolved Ce to <= 3 mg Ce/L (detection limit) was observed. The initial dissolved Ce concentration was 24 mg Ce/L. - Ce was present in the precipitate at the bottom of the vessels.
Results with reference substance (positive control):
- EC50(24h) = 1.6 mg/L
Reported statistics and error estimates:
none
Validity criteria fulfilled:
yes
Conclusions:
This study reported a 48-h EC50 > 100 mg/L (test substance) for Daphnia magna in a limit test. Dissolved Ce analysis indicates that Ce precipitates and disappears from the solution. This should be kept in mind when using the results of the study.

Description of key information

The key study of Hefner (2014b), in which Daphnia magna was exposed for 48 h to cerium ammonium nitrate, yielded a 48-h EC50 of > 26 mg Ce/L (corresponding to > 100 mg (NH4)2Ce(NO3)6/L).

Key value for chemical safety assessment

Additional information

Two studies were included in this dossier as containing relevant information on this endpoint. However, one of the studies (Bazin, 1995) was used as supporting information in the evaluation because the measurements of dissolved cerium were done in test vessels without organisms and this at 0 and 24h. No monitoring was done during the actual test. However, the results of this study were in line with the key study of Hefner (2014b).

The new study performed by Hefner (2014b) was a 48-h study with Daphnia magna in which anhydrous (NH4)2Ce(NO3)6 was used as test item and dissolved cerium concentrations were analytically monitored during the test. The 48-h EC50 was calculated based on nominal concentrations and was > 26 mg dissolved Ce/L (corresponding to > 100 mg (NH4)2Ce(NO3)6/L). These results are considered as the key values. Cerium ammonium nitrate did not show acute toxic effects to Daphnia magna under the conditions of the test up to the solubility limit of the test item in the test water (loading rate of 101 mg/L).

The supportive study was from Bazin (1995). This study reported a 48-h EC50 > 100 mg (NH4)2Ce(NO3)6/L (corresponding to 25.56 mg Ce/L). Cerium disappeared from solution, with concentrations <= 3 mg Ce/L after 24 h of exposure. Data for recalculation of the EC50 value based on measured concentrations was not available. Therefore the real toxicity may have been substantially underestimated.