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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:
6 March 2018 to 20 April 2018
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)
Version / remarks:
2004
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Deviations:
no
Principles of method if other than guideline:
NA
GLP compliance:
yes (incl. QA statement)
Specific details on test material used for the study:
Identification: Rubidium Oxide
Batch: S19D031
Purity: 45.8 wt% in water
Physical state/Appearance: Pale yellow liquid
Expiry Date: 20 October 2018
Storage Conditions: Room temperature in the dark
Analytical monitoring:
yes
Details on sampling:
Samples were taken from the control and each test group from the bulk test preparation at 0 hours and from the pooled replicates at 48 hours for quantitative analysis. All samples were stored frozen prior to analysis. Duplicate samples were taken at 0 and 48 hours and stored frozen for further analysis if necessary.
Vehicle:
no
Details on test solutions:
The test concentrations to be used in the definitive test were determined by preliminary range-finding tests. In the initial range-finding test Daphnia magna were exposed to a series of nominal test concentrations of 0.10, 1.0, 10 and 100 mg a.i./L. A nominal amount of test item (109 mg) was dissolved in test water and the volume adjusted to 500 mL to give the 100 mg a.i./L test concentration from which a series of dilutions was made to give further test concentrations of 0.10, 1.0 and 10 mg a.i./L. Each prepared concentration was inverted several times to ensure adequate mixing and homogeneity. In the range-finding test five daphnids were placed in each test and control vessel and maintained at water temperature at 18 to 22°C with a maximum deviation of ±1 °C with a photoperiod of 16 hours light and 8 hours darkness for a period of 48 hours with 20 minute dawn and dusk transition periods. Two replicate test and control vessels were prepared. Each 150 mL test and control vessel contained 100 mL of test media and was covered to reduce evaporation. After 24 and 48 hours the number of immobilized daphnids were recorded. In the initial range-finding test the pH reading of the 100 mg a.i./L test concentration was outside the range specified in the study plan (pH 6 to 9), therefore a second range-finding test was conducted. In the second range-finding test, the pH of the 100 mg a.i./L test concentration was adjusted using 1.0 M HCl and 0.1 M NaOH, to match the pH of the dilution water. The exposure conditions for the second range-finding test were as described in the initial range-finding test.

Based on the results of the range-finding tests the following test concentrations were assigned to the initial test: 1.0, 1.8, 3.2, 5.6 and 10 mg a.i./L. However, the results of the initial test showed no immobilization at the test concentrations of 3.2, 5.6 and 10 mg a.i./L, 5% immobilization at 1.0 mg a.i./L and 20% immobilization at 1.8 mg a.i./L, therefore, a definitive test was performed using nominal test concentrations of 2.6, 6.4, 16, 40 and 100 mg a.i./L for a period of 48 hours. A nominal amount of test item (436 mg) was dissolved in test water and the volume adjusted to 2000 mL to give the 100 mg a.i./L test concentration. The pH of the stock solution was checked and adjusted using 1.0 M HCl, to match the pH of the dilution water. A series of dilutions was made to give further test concentrations of 2.6, 6.4, 16, 40 mg a.i./L. The concentration and stability of the test item in the test preparations were verified by chemical analysis at 0 and 48 hours.
Test organisms (species):
Daphnia magna
Details on test organisms:
Tests were performed using first instar Daphnia magna derived from in-house laboratory cultures. Adult daphnids were maintained in 150 mL glass beakers containing 100 mL Elendt M7 medium in a temperature controlled room maintaining the water temperature at 18 to 22°C. The lighting cycle was controlled to give a 16 hours light and 8 hours darkness cycle with 20 minute dawn and dusk transition periods. Each culture was fed daily with a mixture of algal suspension (Desmodesmus subspicatus) and Tetramin® flake food suspension. Culture conditions ensured that reproduction was by parthenogenesis. Gravid adults were isolated the day before initiation of the test, such that the young daphnids produced overnight were less than 24 hours old. These young were removed from the cultures and used for testing. The diet and diluent water are considered not to contain any contaminant that would affect the integrity or outcome of the study.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Remarks on exposure duration:
Standard duration
Post exposure observation period:
None, not a standard test requirement
Hardness:
250 mg/L CaCO3
Test temperature:
21°C to 22°C
pH:
7.8 ± 0.1
Dissolved oxygen:
7.6 to 8.9 mgO2/L
Salinity:
NA
Conductivity:
NA
Nominal and measured concentrations:
Nominal test concentrations: 2.6, 6.4, 16, 40 and 100 mg a.i./L (based on two range finding tests)
Details on test conditions:
Exposure Conditions:
In the definitive test 150 mL glass beakers containing approximately 100 mL of test preparation were used. At the start of the test five daphnids were placed in each test and control vessel at random, in the test preparations. Four replicate test and control vessels were prepared. The test vessels were then covered to reduce evaporation and maintained in a temperature controlled room maintaining the water temperature at 18 to 22 °C with a maximum deviation of ±1 °C with a photoperiod of 16 hours light (between 200 and 1200 Lux) and 8 hours darkness with 20 minute dawn and dusk transition periods. The daphnids were not individually identified, received no food during exposure and the test vessels were not aerated. The control group was maintained under identical conditions but not exposed to the test item.

Test Organism Observations:
Any immobilization or adverse reactions to exposure were recorded at 24 and 48 hours after the start of exposure. The criterion of effect used was that daphnia were considered to be immobilized if they were unable to swim within 15 seconds after gentle agitation.

Water Quality Criteria:
Water temperature was recorded daily throughout the test. Dissolved oxygen concentrations and pH were recorded at the start and termination of the test. The pH and dissolved oxygen concentration were measured using a Hach Flexi handheld meter whilst the temperature was measured using a Hanna Instruments HI 93510 digital thermometer. The light intensity during the light period was measured using an ATP Instrumentation Lux meter. The appearance of the test media was recorded daily.
Reference substance (positive control):
yes
Remarks:
Potassium dichromate
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
29 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
16 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
LOEC
Effect conc.:
40 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
mobility
Details on results:
Chemical analysis of the test preparations at 0 and 48 hours showed all measured concentrations were within 20% of the nominal concentrations, with the exception of the 6.4 mg a.i./L 48-Hour old sample which was 77% of nominal. Given that there was no effect on the exposed daphnia at 16 mg a.i./L the slightly low measured concentration at 6.4 mg a.i./L was considered not to adversely affect the results of the test, therefore nominal concentrations were used in calculations of the results.
Results with reference substance (positive control):
The positive control used potassium dichromate as the reference item at concentrations of 0.32, 0.56, 1.0, 1.8 and 3.2 mg a.i./L, with exposure conditions being similar to those in the definitive test. Analysis of the immobilization data was carried out using the Trimmed Spearman-Karber method at 24 hours and the Binomial Distribution method at 48 hours. All statistical analysis was carried out using the ToxRat Professional computer software package with results based on the nominal test concentrations and resulted in EC50 values of 0.79 mg a.i./L and 0.75 mg a.i./L after 24 hour and 48 hour exposures, respectively. Resulting NOEC and LOEC values were 0.56 mg a.i./L and 1.0 mg a.i./L respectively after both 24 hour and 48 hour exposures. The results from the positive control were within the normal range for this reference item.
Reported statistics and error estimates:
An estimate of the EC50, LOEC and NOEC values at 24 hours were given by inspection of the immobilization data.

The EC50 value and associated confidence limits at 48 hours and the slope of the response curve and its standard error were calculated by Probit analysis using Linear Maximum-Likelihood regression. The LOEC and the NOEC at 48 hours were calculated using the Step-down Cochran-Armitage Test. All results were calculated using the ToxRat Professional computer software package (TOXRAT).

Cumulative Immobilization Data and Observations in the Definitive Test

Nominal Concentration (mg a.i./L)

Cumulative Immobilized Daphnia (Initial Population: 5 Per Replicate)

Observations

Rep1

Rep2

Rep3

Rep4

Total

%

Rep1

Rep2

Rep3

Rep4

 

24 hours

Control

0

0

0

0

0

0

5 N

5 N

5 N

5 N

2.6

0

0

0

0

0

0

5 N

5 N

5 N

5 N

6.4

0

0

0

0

0

0

5 N

5 N

5 N

5 N

16

0

0

0

0

0

0

5 N

5 N

5 N

5 N

40

0

0

0

0

0

0

5 N

5 N

5 N

5 N

100

0

0

0

0

0

0

5 N

5 N

5 N

5 N

 

48 hours

Control

0

0

0

0

0

0

5 N

5 N

5 N

5 N

2.6

1

0

0

0

1

5

4 N

5 N

5 N

5 N

6.4

2

0

0

0

2

10

3 N

5 N

5 N

5 N

16

0

0

0

0

0

0

5 N

5 N

5 N

5 N

40

4

2

4

3

13

65

1 N

3 N

1 N

2 N

100

5

5

5

5

20

100

AI

AI

AI

AI

Rep = Replicate

N = Normal

AI = All daphnia immobilized

Validity criteria fulfilled:
yes
Remarks:
None of the control daphnids showed immobilisation or other signs of disease or stress and that the oxygen concentration at the end of the test was equal to or greater than 3 mg a.i./L in the control and test vessels.
Conclusions:
Exposure of freshwater invertebrate Daphnia magna to the test item (rubidium oxide) for 48 hours resulted in a nominal EC50 value of 29 mg a.i./L based on immobilisation.
Executive summary:

The short-term toxicity of Rubidium Oxide to the freshwater aquatic invertebrate, Daphnia magna, was assessed via an OECD Guideline 202 (‘Daphnia sp. Acute Immobilisation Test’) study. The study was performed at a GLP accredited laboratory, fulfilled the validity criteria described by the guideline and is well documented and is therefore considered to be reliable.  

  

Exposures of D. magna to the test item resulted in a nominal 48 hour EC50 value of 29 mg a.i./L based on immobilisation. The resulting 48 hour NOEC and LOEC values were determined to be 16 mg a.i./L and 40 mg a.i./L, respectively.  

Description of key information

The short-term toxicity of Rubidium Oxide to the freshwater aquatic invertebrate, Daphnia magna, was assessed via an OECD Guideline 202 (‘Daphnia sp. Acute Immobilisation Test’) study. The study was performed at a GLP accredited laboratory, met the validity criteria prescribed by the guideline and is well documented and is therefore considered to be reliable.  

  

Exposures of D. magna to the test item resulted in a nominal 48 hour EC50 value of 29 mg a.i./L based on immobilisation. The resulting 48 hour NOEC and LOEC values were determined to be 16 mg a.i./L and 40 mg a.i./L, respectively.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Effect concentration:
29 mg/L

Additional information