Registration Dossier

Ecotoxicological information

Short-term toxicity to aquatic invertebrates

Currently viewing:

Administrative data

Link to relevant study record(s)

Reference
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
25 Apr - 30 Jul 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Version / remarks:
2004
Deviations:
no
Qualifier:
according to
Guideline:
other: OECD No. 23
Version / remarks:
2000
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
Staatliches Gewerbeaufsichtsamt Hildesheim, Germany (03 Jan 2017)
Analytical monitoring:
yes
Remarks:
GC-MS
Details on sampling:
- Concentrations: All concentration levels and the control in fresh (0 and 24 h) and aged (24 and 48 h) media.
- Sampling method: At the start of the exposure and at renewal (0 and 24 h), samples of the fresh media were taken after preparatio of all test item concentrations. At renewal and at the end of the exposure (24 and 48 h), samples of the 24 h old media were taken directly from the test vessels and analyzed.
- Sample storage conditions before analysis: All samples were stored at 6 ± 2 °C until the start of analysis, if necessary. Prepared samples were stored in the autosampler at room temperature until analysis.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: A stock solution of 2.50 mg/L in dilution water was prepared by stirring with a magnetic stirrer at approximately 1100 rpm for approximately 1 h at room temperature. Five concentration levels of the test item in a geometric series with a separation factor of 2 were prepared by dilution of the stock solution with dilution water. - Controls: Dilution water without test item incubated under the same conditions as the test groups.
- Evidence of undissolved material: All concentration levels were visually clear throughout the exposure period.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: water flea
- Strain/clone: Clone 5
- Breeding: Daphnia were bred by the testing facility in glass vessels ( 2 - 3 L capacity) with approximately 1.8 L culture medium, at 20 ± 2 °C, in an incubator, with 16 h illumination and a light intensity of max. 1500 lux. The culture daphnids were fed at least 5 times per week ad libitum with a mix of unicellular green algae, e.g. Pseudokirchneriella subcapitata and Desmodesmus subspicatus, with an algae cell density of > 10^6 cells/mL. The algae were cultured at the test facility.
- Age of daphnids at test start: < 24 h
- Feeding during test: No

METHOD FOR PREPARATION AND COLLECTION OF EARLY INSTARS OR OTHER LIFE STAGES:
Juvenile daphnids were removed from the culture vessels at the latest 24 h before the start of the exposure and discarded. The juveniles born within the following period of max. 24 h preceding the exposure were used for the test. No first brood progeny was used for the test.
Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
169 mg CaCO3/L (0 h)
180 mg CaCO3/L (24 h)
Test temperature:
20.9 °C
pH:
Control: 8.82 (0 h), 8.14 (48 h)
Treatments: 8.75 - 8.94 (0 h), 8.39 - 8.50 (48 h)
Dissolved oxygen:
Control: 9.00 mg/L (0 h), 8.82 mg/L (48 h)
Treatments: 8.78 - 8.81 (0 h), 8.44 - 8.65 (48 h)
Conductivity:
430 µS/cm (0 h)
458 µS/cm (48 h)
Nominal and measured concentrations:
Control, 0.156, 0.313, 0.625, 1.25, and 2.50 mg/L (nominal)
< LOQ , 0.0159, 0.0244, 0.0439, 0.738, and 1.72 mg/L (geometric mean measured)
Details on test conditions:
TEST SYSTEM
- Test vessel: Sealed glass flasks (4.5 I.D. x 9.5 cm H) with screw top were used and filled up to the top with approximiately 130 mL test solution, no headspace
- Type: Closed
- Renewal rate of test solution: Semi-static test with a renewal of test solutions after 24 h
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Conductivity: 430 µS/cm (0 h), 458 µS/cm (24 h)
- Culture medium different from test medium: Culture medium same as test medium
- Intervals of water quality measurement: Water quality was measured at test start (0 h) and at the renewal of the test solutions (24 h).

OTHER TEST CONDITIONS
- Photoperiod: 16 h ligh / 8 h dark
- Light intensity: Diffuse light, max. 1500 lux

EFFECT PARAMETERS MEASURED:
- Immobilization: after 24 and 48 h

RANGE-FINDING STUDY
- Test concentrations: 0.025, 0.25, and 2.5 mg/L (nominal)
- Results used to determine the conditions for the definitive study: Yes, 95% immobilization observed at 2.5 mg/L after 48 h.
Reference substance (positive control):
yes
Remarks:
Potassium dichromate
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
0.044 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95% confidence limits: 0.0277 to > 1.72 mg/L
Details on results:
- Behavioural abnormalities: None
- Mortality of control: 0%
- Any observations that might cause a difference between measured and nominal values: All tested concentration levels were visually clear throughout the exposure period.
- Effect concentrations exceeding solubility of substance in test medium: No
Results with reference substance (positive control):
- Results with reference substance valid? Yes, results were within the valid range.
- Relevant effect levels: EC50 (48 h) = 1.31 mg/L, 95% confidence limits 1.05 - 1.72 mg/L
- Other: Exposure phase 11 - 12 Jul 2017
Reported statistics and error estimates:
The EC10/EC50 values after 24 and 48 h were calculated by sigmoidal dose-response regression with the software GraphPad Prism. The respective 95% confidence limits were calculated from the standard error and the t-distribution. All calculations were carried out from the best-fit values with the software GraphPad Prism.

VALIDITY CRITERIA

 The study fulfilled the validity criteria defined by the guideline (Table 1).

 

Table 1: Validity criteria

Criterion from the guideline

Outcome

Validity criterion fulfilled

In the control, including the control containing the solubilising agent, not more that 10% of the daphnids should have been immobilized.

No immobilization in the control.

Yes

The dissolved oxygen concentration at the end of the test should be ≥ 3 mg/L in control and test vessels.

The dissolved oxygen in the 24 h old media was ≥ 7.58 mg/L.

Yes

 

 

ANALYTICAL RESULTS

 The measured concentrations of the test item in fresh media at the start of the exposure and at renewal (0 and 24 h) were in the range of 50 to 92% of the nominal test item concentrations. The measured concentrations in old media at renewal and at the end of the test (24 and 48 h) were in the range of 0 to 65% of the nominal test item concentrations (Table 2).

 

Table 2. Measured concentrations of the test item during the definitive test.

Sampling date

0 h

Start of the exposure interval

24 h

End of the exposure interval

24 h

Start of the exposure interval

48 h

End of the exposure interval

Geometric mean measured test item concentration [mg/L]

Nominal test item concentration [mg/L]

Measured concentration [mg/L]

%

Measured concentration [mg/L]

%

Measured concentration [mg/L]

%

Measured concentration [mg/L]

%

2.50

2.01

81

1.61

65

2.18

87

1.23

49

1.72

1.25

1.02

81

0.512

41

1.09

88

0.519

42

0.738

0.625

0.498

80

0.005172)

<1

0.576

92

< LOQ1)

< LOQ1)

< LOQ1)

0.0439

0.313

0.208

66

< LOQ1)2)

< LOQ1)2)

0.275

88

0.0244

0.156

0.0781

50

0.132

85

0.0159

Control

< LOQ

< LOQ

< LOQ3)

< LOQ2)

-

Measured concentration= measured concentration of the test item, enrichment and dilution factors taken into account

%= percentage of the initially measured concentration of the test item

LOQ= limit of quantification of the analytical method (0.005 mg test item/L)

1)= concentration < LOQ, therefore taken with ½ LOQ (0.0025 mg test item/L into account for calculation of the geometric mean

2)= extract of 2017-07-27 reanalyzed on 2017-07-28, undiluted

3)= reanalyzed on 2017-07-30

 

 

BIOLOGICAL RESULTS

The effect concentration (EC10, EC50 and EC100) were based on the geometric mean measured concentrations of the test item.

 

Table 3. Immobilization Rates after 24 and 48 hours of Exposure in the Definitive Test.

Geometric mean measured concentration of the test item [mg/L]

Immobilization [%]

24 h

48 h

Replicates

Replicates

 

1

2

3

4

MV

1

2

3

4

MV

1.72

20

100

80

80

70

100

100

100

100

100

0.738

20

0

0

0

5

20

80

40

20

40

0.0439

80

60

40

0

45

80

80

40

0

50

0.0244

0

0

0

0

0

0

20

20

20

15

0.0159

0

0

0

0

0

0

0

0

0

0

Control

0

0

0

0

0

0

0

0

0

0

MV = mean value

 

Validity criteria fulfilled:
yes
Remarks:
For further details please refer to “Any other information on results incl. tables”.
Conclusions:
Based on the test outcome the EC50 (48 h) for Daphnia magna was 0.0436 mg/L (geometric mean measured, OECD 202).

Description of key information

EC50 (48 h) = 0.0436 mg/L (geom. mean measured, OECD 202)

Key value for chemical safety assessment

EC50/LC50 for freshwater invertebrates:
0.044 mg/L

Additional information

There is one study available, in which the short-term toxicity of the substance toward aquatic invertebrates was assessed according to the OECD guideline 202 and GLP.

In a semi-static test design, Daphnia magna was exposed to five concentrations in a geometric series with a dilution factor of 2 ranging from 2.50 to 0.156 mg/L test item for 48 h in sealed glass flasks without headspace. The test solutions were renewed after 24 h and the test item concentrations in the test vessels were analytically verified by GC-MS in fresh (0 and 24 h) and old (24 and 48 h) media.

The measured concentrations of the test item in fresh media at the start of the exposure and at renewal (0 and 24 h) were in the range of 50 to 92% of the nominal test item concentrations. The measured concentrations in old media at renewal and at the end of the test (24 and 48 h) were in the range of 0 to 65% of the nominal test item concentrations. Therefore, the effect concentrations were based on the geometric mean measured concentrations.

After 48 h, immobilization was observed and an EC50 (48 h) of 0.0436 mg/L was determined.