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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:
09 November 2017 - 18 December 2017
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
GLP compliance:
yes (incl. QA statement)
Analytical monitoring:
yes
Details on sampling:
- Concentrations: 100, 45.5, 20.7, 9.38 and 4.27 mg/L (nominal test item concentrations)
- Sampling method: At the start of the exposure and at renewal (0 and 24 hours), samples of the fresh media were taken after preparation of all test item concentrations and analyzed. At renewal and at the end of the exposure (24 and 48 hours), samples of the 24-hours 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 the 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 (100 mg/L of the test item were weighed out) was prepared with dilution water prior to the start of the exposure (at 0 hours) and prior to the renewal of the test solutions (at 24 hours). The stock solution was mixed thoroughly by manual agitation until the test item was completely dissolved. 5 test item concentrations (4.27, 9.39, 20.7, 45.5 and 100 mg/L) in a geometric series with a separation factor of 2.2, prepared by dilution of the stock solution 100 mg/L with dilution water.
- Controls: Dilution water without test item incubated under the same conditions as the test groups.
- Evidence of undissolved material: no
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: water flea
- Strain/clone: Straus (Clone 5)
- Source: Origin: Institut für Wasser-, Boden- und Lufthygiene (WaBoLu), 14195 Berlin, Germany, Breeder: Noack Laboratorien GmbH, Käthe-Paulus-Str. 1, 31157 Sarstedt, Germany
- Age of parental stock: less than 24 hours old
- Feeding during test: no
- Food type: mix of unicellular green algae, e.g. Pseudokirchneriella subcapitata and Desmodesmus subspicatus, with an algae cell density of > 106 cells/mL
- Frequency: at least 5 times per week ad libitum

ACCLIMATION
- Acclimation period: not necessary, because the composition of the dilution water was equivalent to the culture medium
Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
160 to 180 mg CaCO3/L
Test temperature:
18 - 20 °C
Dissolved oxygen:
≥ 7.60 mg/L
Nominal and measured concentrations:
- nominal concentrations: 100, 45.5, 20.7, 9.38 and 4.27 mg/L
- measured concentrations: new media ( 0 and 24 h): 98 to 103 % of the nominal values, old media: 24 and 48 h): 88 to 98 % of the nominal values (see table 1)
Details on test conditions:
TEST SYSTEM
- Test vessel: Sealed glass flasks with screw
- Type: closed
- Material, size, headspace, fill volume: glass 4.5 (ID) x 9.5 (H) cm, without headspace, approximately 130 mL
- 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: Culture Medium Elendt M4 according to OECD 202, Annex 3 (2004)
- Culture medium different from test medium: no
- Intervals of water quality measurement: Prior to the start of the exposure (0 hours) and the renewal of the test solutions (24 hours), the water quality parameters (i.e. pH value, dissolved oxygen concentration, temperature, conductivity and total hardness) of the dilution water were measured.

OTHER TEST CONDITIONS
- Photoperiod: 16/8 hours light/dark cycle
- Light intensity: max. 1500 lx

RANGE-FINDING STUDY
- Test concentrations: 100, 10 and 1 mg/L (nominal test item concentration)
- Results used to determine the conditions for the definitive study: yes
Reference substance (positive control):
yes
Remarks:
Potassium dichromate
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
9.55 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
EC10
Effect conc.:
< 4.27 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
EC100
Effect conc.:
45.5 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: none
- Effect concentrations exceeding solubility of substance in test medium: none
Results with reference substance (positive control):
- Results with reference substance valid? yes
- EC50 (24 h): 1.64 mg/L

Table 1: Measured Concentrations of the Test Itemduring the Definitive Test

Sampling
date

Fresh media,

0 hours

Old media,

24 hours

Fresh media,

24 hours

Old media,

48 hours

Nominal test item

concentration

[mg/L]

test item

Meas. conc.

[mg/L]

%

Meas. conc.

[mg/L]

%

Meas. conc.

[mg/L]

%

Meas. conc.

[mg/L]

%

100

100

100

96.6

97

100

100

96.8

97

 45.5

46.7

103

44.2

97

45.6

100

44.6

98

  20.7

21.1

102

20.2

97

21.0

102

19.4

93

    9.39

9.39

100

8.93

95

9.38

100

8.55

91

    4.27

4.34

102

3.96

93

4.21

98

3.77

88

Control

< LOQ

< LOQ

< LOQ

< LOQ

Meas. conc.    = measured concentration of the test item, dilution factors taken into account, mean value of 2 injections

%                     = percentage of the nominal concentration of the test item

LOQ                = limit of quantification of the analytical method (2.50 mg test item/L)

 

Table 2: Immobilization Rates after 24 and 48 hours of Exposure in the Definitive Test (n = 20, divided into 4 replicates with 5 daphnids each)

Nominal test item

concentration

[mg/L]

IMMOBILIZATION [%]

24 hours

48 hours

Replicates

Replicates

1

2

3

4

MV

1

2

3

4

MV

100

100

80

80

100

90

100

100

100

100

100

45.5

60

60

40

40

50

100

100

100

100

100

20.7

20

20

20

0

15

80

60

60

20

55

9.39

0

20

20

20

15

60

60

60

60

60

4.27

0

0

0

0

0

40

20

0

20

20

Control

0

20

0

0

5

0

40

0

0

10

Table 3: Absolute Numbers of immobile Daphnids after 24 and 48 hours of Exposure in the Definitive Test (n = 20, divided into 4 replicates with 5 daphnids each)

Nominal test item

concentration

[mg/L]

Number of immobile Daphnids / Total number of Daphnids

24 h

48 h

Replicates

Replicates

1

2

3

4

MV

1

2

3

4

MV

100

5 / 5

4 / 5

4 / 5

5 / 5

18 / 20

5 / 5

5 / 5

5 / 5

5 / 5

20 / 20

 45.5

3 / 5

3 / 5

2 / 5

2 / 5

10 / 20

5 / 5

5 / 5

5 / 5

5 / 5

20 / 20

  20.7

1 / 5

1 / 5

1 / 5

0 / 5

3 / 20

4 / 5

3 / 5

3 / 5

1 / 5

11 / 20

    9.39

0 / 5

1 / 5

1 / 5

1 / 5

3 / 20

3 / 5

3 / 5

3 / 5

3 / 5

12 / 20

    4.27

0 / 5

0 / 5

0 / 5

0 / 5

0 / 20

2 / 5

1 / 5

0 / 5

1 / 5

4 / 20

Control

0 / 5

1 / 5

0 / 5

0 / 5

1 / 20

0 / 5

2 / 5

0 / 5

0 / 5

2 / 20

 

Validity criteria fulfilled:
yes
Conclusions:
The test item had acute toxic effects on Daphnia magna in a 48-hour semi-static test. The 48-hour EC50 was calculated to be 9.55 mg/L.
Executive summary:

In the acute immobilization test with Daphnia magna (STRAUS), the effects of the test item were determined according to OECD 202 (2004).

The study was conducted in a closed system (sealed glass flasks) without headspace under semi-static conditions over a period of 48 hours with the stock solution of the test item (100 mg/L) and further four dilution levels (nominal: 4.27 – 45.5 mg/L) prepared from the stock solution in a geometric series with a separation factor of 2.2. Twenty daphnids (divided into 4 replicates with 5 daphnids each) were exposed to each concentration level and the control. The concentrations of the test item were analytically verified via HPLC-DAD in fresh media at the start of the exposure and at renewal (0 and 24 hours) and in old media at renewal and at the end of the test (24 and 48 hours) in all concentration levels and the control. The measured concentrations in fresh media at the start of the test and at renewal (0 and 24 h) were between 98 and 103% of the nominal values. The measured concentrations in old media at renewal and at the end of the test (24 and 48 hours) were in the range of 88 to 98% of the nominal concentrations.

As the measured concentrations were within the range of ± 20% of the nominal values, the effect concentrations are based on the nominal concentrations of the test item. After 48 h, the determined EC50 value was 9.55 mg/L.

Description of key information

The 48-hour EC50 was calculated to be 9.55 mg/L.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Effect concentration:
9.55 mg/L

Additional information

In the acute immobilization test withDaphnia magna(STRAUS), the effects of the test item were determined according to OECD 202 (2004).

The study was conducted in a closed system (sealed glass flasks) without headspace under semi-static conditions over a period of 48 hours with the stock solution of the test item (100 mg/L) and further four dilution levels (nominal: 4.27 – 45.5 mg/L) prepared from the stock solution in a geometric series with a separation factor of 2.2. Twenty daphnids (divided into 4 replicates with 5 daphnids each) were exposed to each concentration level and the control. The concentrations of the test item were analytically verified via HPLC-DAD in fresh media at the start of the exposure and at renewal (0 and 24 hours) and in old media at renewal and at the end of the test (24 and 48 hours) in all concentration levels and the control. The measured concentrations in fresh media at the start of the test and at renewal (0 and 24 h) were between 98 and 103% of the nominal values. The measured concentrations in old media at renewal and at the end of the test (24 and 48 hours) were in the range of 88 to 98% of the nominal concentrations.

As the measured concentrations were within the range of ± 20% of the nominal values, the effect concentrations are based on the nominal concentrations of the test item. After 48 h, the determined EC50 value was 9.55 mg/L