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Ecotoxicological information

Short-term toxicity to aquatic invertebrates

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Administrative data

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
24 January 2012 to 2 February 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. The study report was conclusive, done to valid guidelines and the study was conducted under GLP conditions.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report Date:
2012

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 850.1010 (Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids)
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
other: solid (unspecified)
Details on test material:
- Name of test material (as cited in study report): Cobalt Lithium Manganese Nickel oxide
- Appearance: black solid
- Empirical formula: Li (Co 0.33 Mn 0.33 Ni 0.33) O₂
- Homogeneity: homogeneous
- Storage condition of test material: ambient; under nitrogen
- Storage stability: the stability of the test material under the storage conditions specified was guaranteed by the Sponsor for the duration of the test.

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
At the start of the exposure (0 hours) samples were taken from additionally prepared test vessels without daphnids and at the end of the exposure (48 hours) samples from all test vessels with mobile daphnids were analysed.
The samples were stored frozen until analysis.

Test solutions

Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
Since the test material was considered to be poorly soluble in water, the test solutions were prepared following general guidance recommended by OECD 23 and US EPA OPPTS 850.1000 in order to achieve a saturated solution of the test material.
200.25 mg of test material was mixed with 2 L test medium (M4). The mixture was stirred on a magnetic stir plate for about 7 days (as recommended in OECD 29) at 20 ± 2 °C. Undissolved test material was distributed in the test solution vessel. To avoid any physical injury to the test organisms and ensure testing of only the aqueous dissolved fraction of the test material, undissolved material was removed prior to exposure, as recommended in OECD 23, by filtration through a cellulose acetate filter (pore width 0.2 µm, Whatman OE66). The filter was conditioned with test solution before collecting the final solution. The exposure was started after separation of undissolved material.
After separation, the test solution was considered to be a water accommodated fraction (WAF). The test solution appeared clear and colourless after filtration.

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Strain: Daphnia magna STRAUS 1820
- Age at study initiation: < 24 hours (first instar); at least 3rd brood progeny
- Method of breeding: by parthenogenesis. Brood stock are kept in mass cultures consisting of 20-30 parthenogenetically reproducing females for a minimum of 4 weeks and fed live unicellular green algae; they are cultured continuously. All individuals in each mass culture originate from a single female and are therefore genetic clones. After approximately 14 days the adults have produced at least 3 broods and the live young can be used in tests.
- Feeding during test: no
- Health of stock: only young from healthy cultures without signs of stress were used for testing.

ACCLIMATION
The daphnia are cultured under identical conditions as the test including test media (Elendt M4), water quality and temperature (20 ± 1 °C).

Study design

Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
48 h

Test conditions

Hardness:
2.40 mmol/L
Test temperature:
19.8 - 20.2 °C
pH:
8.0 - 8.3
Dissolved oxygen:
5.4 - 6.0 mg/L (after 0 hours); 8.9 - 9.1 mg/L (after 48 hours)
Nominal and measured concentrations:
0 (medium control) and 100 mg/L (nominal)
Details on test conditions:
TEST SYSTEM
- Test vessel: 250 mL glass beakers
- Fill volume: 100 mL
- Aeration: no
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Test medium / dilution water: Synthetic freshwater (Elendt M4)
- Total organic carbon: < 2 mg/L
- Total hardness: 2.20 - 3.20 mmol/L
- Acid capacity up to pH 4.3: 0.80 - 1.00 mmol/L
- pH: 7.5 - 8.5
- Ca:Mg ratio: about 4 : 1
- Conductivity: 550 - 650 μS/cm
To ensure that dissolved oxygen levels are at or above 3 mg/L during the test, the Elendt M4 medium is aerated for approximately 24 hours prior to use.
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 hours light / 8 hours dark
- Light intensity: 161 - 664 lux at a wavelength of 400 - 750 nm

EFFECT PARAMETERS MEASURED
Each test vessel was visually inspected for immobilised daphnids at 0, 24 and 48 hours; in addition, any abnormal behaviour or appearance was documented.

PHYSICAL PARAMETERS MEASURED
Throughout the test, the appearance of the test solutions and dissolution behaviour of the test material was observed and recorded daily. The chemical and physical parameters of the test medium (TOC, total hardness, acid capacity, pH and conductivity) were measured after aeration and prior to use in the test. Dissolved oxygen and pH were measured at the start and at the end of the exposure period in both the test solution and control. In addition, temperature was measured continuously during the whole exposure period in a separate vessel filled with water proximal to the test vessels.

TEST CONCENTRATIONS
- Range finding study: In a preliminary test a concentration of 100 mg/L (loading rate), filtered to remove undissolved material, caused no immobilisation or symptoms after 48 hours. Accordingly, a limit test was conducted using the highest recommended test concentration of test material.
Reference substance (positive control):
yes
Remarks:
potassium dichromate

Results and discussion

Effect concentrations
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
None of the daphnia were observed to be immobile at any observation point during the study. No additional adverse effects or abnormal behaviour were observed in any of the test groups.
The test organisms were exposed to a saturated solution of all soluble components of the test material at the given loading rate and, therefore, the effect concentration was based on the loading rate.
All of the test solutions appeared to be visibly clear after preparation and remained so after 24 and 48 hours of exposure in the test. No undissolved test material was visible and there were no other remarkable observations.

VALIDITY CRITERIA
The test was fully compliant with the following validity criteria:
- ≤ 10 % immobilisation in the control.
- Oxygen concentration > 3 mg/L in control and test vessels.
Results with reference substance (positive control):
- Results with reference substance valid? Yes
- 24-hour EC50: 0.70 mg/L
This result was within the range of 0.6 - 2.1 mg/L (ISO 6341) and indicates that the daphnia culture used in the study responded normally to toxic stress.

Any other information on results incl. tables

Analytical Determinations

The analytically determined concentrations of Co and Ni in the test material solutions were:

Co content: 19.9 g/100 g

Ni content: 19.7 g/100 g

 

Table 1: Analytically measured concentrations

Nominal loading rate (mg/L)

0 hours

48 hours

Co

(mg/L)

Ni

(mg/L)

Test material

(mg/L)

Co

(mg/L)

Ni

(mg/L)

Test material

(mg/L)

0

< 0.03

< 0.03

< 0.2

< 0.03

< 0.03

< 0.2

100

< 0.03

0.03

< 0.2

< 0.03

0.03

< 0.2

 

Cobalt concentrations were below the LOQ at the start and end of exposure. Ni concentrations were stable over the exposure period at 0.03 mg/L. However, the estimated equivalent amount of test material in solution was < 0.2 mg/L. Nevertheless, the test solutions were considered saturated with all soluble components of the test material, in test media and under test conditions.

All reasonable efforts were used to generate a saturated solution of the test material in test media following OECD guideline 23. Since the test material is a mixed metal oxide, the test solution is considered a water accommodated fraction (WAF).

Table 2: Number of immobilised Daphnids at each observation period

Nominal loading rate (mg/L)

Replicate 1

Replicate 2

Replicate 3

Replicate 4

0 h

24 h

48 h

0 h

24 h

48 h

0 h

24 h

48 h

0 h

24 h

48 h

24 h

48 h

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

100

0

0

0

0

0

0

0

0

0

0

0

0

0

0

Number per replicate: 5

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
Under the conditions of the study, the 48-hour EL50 of the test material to freshwater invertebrates was determined to be in excess of 100 mg/L based on the loading rate of the test material in the test media. The results of the study demonstrate that the test material is not acutely toxic to daphnia up to the limit of solubility.
Executive summary:

The short term toxicity of the test material to freshwater invertebrates was investigated in a study which was conducted under GLP conditions and in accordance with the standardised guidelines OECD 202, EU Method C.2 and US EPA OPPTS 850.1010.

During the study groups of 20 Daphnia magna STRAUS were each exposed to test material at nominal concentrations of 0 (dilution water control) and 100 mg/L, under static conditions, for 48 hours. Immobilisation, adverse effects and any signs of abnormal behaviour were recorded.

The content of cobalt and nickel in the test solutions was analysed during the study. Cobalt concentrations were below the LOQ at the start and end of exposure. Nickel concentrations were stable over the exposure period at 0.03 mg/L. However, the estimated equivalent amount of test material in solution was < 0.2 mg/L. Nevertheless, the test solutions were considered saturated with all soluble components of the test material, in test media and under test conditions.

Since the daphnia were exposed to a saturated solution of test material at the given loading rate, the effect concentrations were based on the loading rate (as recommended in OECD 23).

Under the conditions of the study, a saturated solution of the test material prepared at a loading rate of 100 mg/L in test media had no observable acute toxic effect on the daphnia with no immobilisation seen. The 48-hour EL50 was therefore determined to be in excess of 100 mg/L. The results of the study demonstrate that the test material is not acutely toxic to daphnia up to the limit of solubility.