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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:
15/1/2013 - 17/1/2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study according to international guideline (OECD guideline 202) under GLP. No deviations from guideline reported.
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Deviations:
no
Principles of method if other than guideline:
Not relevant
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
Not relevant
Analytical monitoring:
yes
Details on sampling:
Not relevant
- Concentrations: 1.0, 3.2, 10, 32 and 100 mg/L WAF
- Sampling method: Samples were taken from the control and each test group (replicates R1 - R4 pooled) at 0 and 48 hours.
- Sample storage conditions before analysis: stored at -20ºC
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Amounts of test item (22.6, 36.2, 56, 76.8 and 240 mg) were each separately added to the surface of 22.6, 11.3, 5.6, 2.4 and 2.4 liters of reconstituted water to give the 1.0, 3.2, 10, 32 and 100 mg/L loading rates respectively. The test item was weighed out under non-actinic light conditions due to the light-sensitive nature of the test item. After the addition of the test item, the vessels were sealed to prevent losses through volatilization and shielded from light. The
reconstituted water was stirred by magnetic stirrer using a stirring rate such that a vortex was formed to give a dimple at the water surface. The stirring was stopped after 4 7 hours and the mixtures allowed to stand for 1 hour. A wide bore glass tube, covered at one end with Nescofilm was submerged into the vessel, sealed end down, to a depth of approximately 5 cm from the bottom of the vessel. A length of Tygon tubing was inserted into the glass tube and pushed
through the Nescofilm seal. Microscopic inspection of the W AFs showed no micro-dispersions or undissolved test item to be present. The aqueous phase or W AF was removed by mid-depth siphoning (the first approximate 75-100 mL discarded) to give the 1.0 , 3.2, 10, 32 and 100 mg/L
loading rate W AFs.
- Controls: Yes, blanks
- Evidence of undissolved material (e.g. precipitate, surface film, etc): At the start of the mixing period the 1.0, 3.2, 10, 32 and 100 mg/L loading rates were observed to be clear colorless water columns with oily globules of test item on the surface. After 47 hours stirring and a 1-Hour standing period the 1.0, 3.2, 10, 32 and 100 mg/L loading rates were observed to remain clear colorless water columns with oily globules of test item on the surface. Microscopic inspection of the W AFs showed no micro-dispersions or undissolved test item to be present. After siphoning and for the duration of the test, the 1.0, 3.2, 10, 32 and 100 mg/L loading rates were observed to be clear, colorless solutions.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia magna
- Source: in-house laboratory strain
- Age at study initiation (mean and range, SD): less than 24 hours old
- Method of breeding: Adult Daphnia were maintained in 150 mL glass beakers containing Elendt M7 medium in a temperature controlled room at 20 °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 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.
- Feeding during test: no

ACCLIMATION
- Acclimation period: 24 hours
- Acclimation conditions (same as test or not): same
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Post exposure observation period:
Not relevant
Hardness:
250 mg/L CaCO3
Test temperature:
20 - 22ºC
pH:
7.6 - 8.1
Dissolved oxygen:
7.5 - 9.3 mg/L (82 - 107% of ASV)
Salinity:
Not relevant
Nominal and measured concentrations:
Nominal loading rates: 1, 3.2, 10, 32 and 100 mg/L
TOC (0h): 5.25, 10.04, 15.46, 21.67 and 25.19 mg/L
TOC (48h): 5.55, 9.59, 12.89, 19.62 and 21.34 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: test vessel
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: Glass, 130 mL filled with 130 mL test medium
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: reconstituted water
- Culture medium different from test medium: No
- Conductivity: <5 uS/cm^-1
- Intervals of water quality measurement: 24 hours

OTHER TEST CONDITIONS
- Photoperiod: 16/8 hours light/dark with 20 minute transition period

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): immobilisation (24 h)

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.2
- Range finding study
- Test concentrations: 1, 10 and 100 mg/L
- Results used to determine the conditions for the definitive study: yes
Reference substance (positive control):
yes
Remarks:
Potassium dichromate
Duration:
24 h
Dose descriptor:
EL50
Effect conc.:
> 17 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95%CI: 10 -32 mg/L
Duration:
48 h
Dose descriptor:
EL50
Effect conc.:
6 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95%CI: 5.4 - 6.7 mg/L
Duration:
24 h
Dose descriptor:
NOELR
Effect conc.:
10 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
48 h
Dose descriptor:
NOELR
Effect conc.:
3.2 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
- Mortality of control: 0%
- Effect concentrations exceeding solubility of substance in test medium: At the start of the mixing period the 1.0, 3.2, 10, 32 and 100 mg/L loading rates were observed to be clear colorless water columns with oily globules of test item on the surface. After 47 hours stirring and a 1-Hour standing period the 1.0, 3.2, 10, 32 and 100 mg/L loading rates were observed to remain clear colorless water columns with oily globules of test item on the surface. Microscopic inspection of the W AFs showed no micro-dispersions or undissolved test item to be present. After siphoning and for the duration of the test, the 1.0, 3.2, 10, 32 and 100 mg/L loading rates were observed to be clear, colorless solutions.
Results with reference substance (positive control):
- Results with reference substance valid? Yes
- 48h-EC50: 0.45 mg/L
Reported statistics and error estimates:
The EL 50 values and associated confidence limits at 48 hours were calculated by the trimmed Spearman-Karber method (Hamilton et al, 1977) using the ToxCalc computer software package (ToxCalc, 1999).When only one partial response is shown the trimmed Spearman-Karber method is appropriate.
If there is no immobilization between 0% and 100% immobilization, then the geometric mean of the highest loading rate showing no lethality and the lowest loading rate showing 100% lethality is calculated. The concentrations resulting in 0% and 100% immobilization will be the 95%
confidence limits.

Cumulative immobility:

Nominal loading rate (mg/L)

Cumulative immobilized Daphnia (initial population: 5 per replicate)

24 hours

48 hours

R1

R2

R3

R4

Total

%

R1

R2

R3

R4

Total

%

Control

0

0

0

0

0

0

0

0

0

0

0

0

1.0

0

0

0

0

0

0

0

0

0

0

0

0

3.2

0

0

0

0

0

0

0

0

0

0

0

0

10

0

0

0

0

0

0

5

4

5

5

19

95

32

5

5

5

5

20

100

5

5

5

5

20

100

100

5

5

5

5

20

100

5

5

5

5

20

100

Graphs are presented below.

Validity criteria fulfilled:
yes
Remarks:
No mortality in controls, physical-chemical parameters within acceptable limits
Conclusions:
The acute toxicity (48h-EL50) of Cymbopogon winterianus towards Daphnia magna is 6.0 mg/L.
Executive summary:

The acute toxicity of Cymbopogon winterianus towards Daphnia magna was investigated according to OECD guideline 202 under GLP. Daphnids were exposed to nominal WAF's of 1.0, 3.2, 10, 32 and 100 mg/L and observed for 48 hours. The 48h-EL50 was found to be 6.0 mg/L based on nominal concentrations.

Description of key information

The 48h-EL50 is 6.0 mg/L in an OECD guideline 202 test with Daphnia magna under GLP (reliability 1).

Key value for chemical safety assessment

EC50/LC50 for freshwater invertebrates:
6 mg/L

Additional information

The acute toxicity of Cymbopogon winterianus towards Daphnia magna was investigated according to OECD guideline 202 under GLP. Daphnids were exposed to nominal WAF's of 1.0, 3.2, 10, 32 and 100 mg/L and observed for 48 hours. The 48h-EL50 was found to be 6.0 mg/L based on nominal concentrations.