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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:
2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Version / remarks:
published in O.J. L 142 (2008)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Test substance name: Acid Brown 235
- Other name: Korostan Brown DGR
- Source and lot/batch No. of test material: 7012/2007
- Expiration date of the lot/batch: May 2018
- Appearance: dark brown powder
Analytical monitoring:
yes
Details on sampling:
The analytical determination of the test substance concentrations was performed at the beginning and at the end of the test. The samples for analysis were taken from the highest (100 mg/L) and the lowest test concentration (10 mg/L). The samples for analysis (0 hours) were prepared at the beginning of the test and immediately delivered in transport box to analytical laboratory. An analysis of samples was started immediately after delivery. The samples for analysis at the end of the test (48 hours) were delivered to analytical laboratory immediately after the end of testing. An analysis of samples was started immediately after delivery. All samples were stored at laboratory temperature.
The analytical results showed that the test substance was stable in dilution water under current test conditions.
Vehicle:
no
Details on test solutions:
For holding and culturing of the daphnia the M4 medium was used (according to Annex 2 of the EU method C.2 and according to Annex 3 of the OECD TG 202).
For test was used dilution water (according to Annex 2 of the EU method C.2 and according to Annex 3 of the OECD TG 202). Dilution water was prepared just before the test by admixing 2.5 mL of stored solutions A, B, C and D into 1 000 mL of deionized water.
A: 117.6 g CaCl2·2H20 in 1L deionized water
B: 49.3 g MgSO4·7H2O in 1L deionized water
C: 25.9 g NaHCO3 in 1L deionized water
D: 2.3 g KCl in 1L deionized water
The dilution water should have a pH 7.8 ± 0.2, values measured in this study were 8.0 in the preliminary test.
The content of sum Ca + Mg ions should have the value of 2.55 ± 0.10 mmol/L, values measured in this study were 2.55 mmol/L in the preliminary test.
Measured values meet the requirements.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia magna
- Strain/clone: Straus
- Source: obtained by aperiodic parthenogenesis from own laboratory breeding
- Feeding during test: no
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
48 h
Test temperature:
20±2°C, with a variation of ± 1°C at each individual test
pH:
7.6 - 8.0
Dissolved oxygen:
8.2 - 8.6 mg/L
Nominal and measured concentrations:
Nominal concentration: 10 mg/L of the test substance
8.33 mg/L (0 h)
8.29 mg/L (48 h)
Nominal concentration: 100 mg/L of the test substance
96.6 mg/L (0 h)
102.1 mg/L (48 h)
Details on test conditions:
TEST SYSTEM
- Test vessel: laboratory beakers
- Aeration: no
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 1

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: daylight
Reference substance (positive control):
yes
Remarks:
potassium dichromate (CAS: 7778-50-9)
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
24 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
EC0
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
24 h
Dose descriptor:
EC0
Effect conc.:
100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
EC100
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
24 h
Dose descriptor:
EC100
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
see Table 1
Results with reference substance (positive control):
RESULTS WITH REFERENCE SUBSTANCE
- Identification of the reference substance: potassium dichromate (CAS: 7778-50-9)
- EC50 = 0.62 mg/L (48 h, Daphnia magna)
- Interlaboratory test data: EC50 = 0.46 - 1.08 mg/L (48 h, Daphnia magna)

Table 1: Immobilisation of daphnia

Nominal concentration

mg/L

100

80

40

20

10

Control

Number
of mobile animals

24 hours

1stseries

10

10

10

10

10

10

Number
of mobile animals

24 hours

2ndseries

10

10

10

10

10

10

Number
of mobile animals

24 hours

total

20

20

20

20

20

20

Immobilisation %

24 hours

0

0

0

0

0

0

Number
of mobile animals

48 hours

1stseries

10

10

10

10

10

10

Number
of mobile animals

48 hours

2ndseries

10

10

10

10

10

10

Number
of mobile animals

48 hours

total

20

20

20

20

20

20

Immobilisation %

48 hours

0

0

0

0

0

0

Validity criteria fulfilled:
yes
Conclusions:
The experiment with the concentration of 100 mg/L included in the preliminary test has been performed using 20 daphnia, therethrough the conditions of the limit test given in guidelines have been fulfilled. It was demonstrated that the EC50 is higher than this concentration.
No immobilisation was observed in the preliminary test. Therefore exact values of EC50 could not be calculated and the values of EC are given in the form of a range. The EC0 and EC100 values are identified by direct observation.
The analytical results from the preliminary test showed, that the test substance, Acid Brown 235, was stable in the dilution water at conditions of the test.
There is stated in the guideline that, if evidence is available to demonstrate that the concentration of the test substance has been satisfactorily maintained within ± 20 percent of the nominal or measured initial concentration throughout the test, the results can be based on nominal or measured initial values. The nominal concentrations were used for all evaluation and results.
Test results:
EC50 > 100 mg/L (Daphnia magna, 24 h) (nominal concentration)
EC50 > 100 mg/L (Daphnia magna, 48 h) (nominal concentration)
EC0 = 100 mg/L (Daphnia magna, 24 h) (nominal concentration)
EC0 = 100 mg/L (Daphnia magna, 48 h) (nominal concentration)
EC100 > 100 mg/L (Daphnia magna, 24 h) (nominal concentration)
EC100 > 100 mg/L (Daphnia magna, 48 h) (nominal concentration)
Executive summary:

The test substance, Acid Brown 235, was tested in acute immobilisation test on Daphnia magna. The test was performed according to method C.2 – Daphnia sp. Acute Immobilisation Test, Council Regulation (EC) No. 440/2008, published in O.J.L 142, 2008.

The preliminary test was performed in a range of the test substance nominal concentrations 10 – 100 mg/L. The test was performed as static. Based on no toxicity of the test substance found in preliminary test, the study was terminated. The experiment with the concentration of 100 mg/L included in the preliminary test has been performed using 20 daphnia, therethrough the conditions of the limit test given in guideline have been fulfilled. It was demonstrated that the EC50 is higher than this concentration.

The samples for analytical determination were taken at the beginning and at the end of the test. The analytical results from the preliminary test showed, that the test substance, Acid Brown 235, was stable in the dilution water at conditions of the test.

There is stated in the guideline that, if evidence is available to demonstrate that the concentration of the test substance has been satisfactorily maintained within ± 20 percent of the nominal or measured initial concentration throughout the test, the results can be based on nominal or measured initial values.

The nominal concentrations were used for all evaluation and results.

Test results:

EC50 > 100 mg/L (Daphnia magna, 24 h) (nominal concentration)

EC50 > 100 mg/L (Daphnia magna, 48 h) (nominal concentration)

EC0 = 100 mg/L (Daphnia magna, 24 h) (nominal concentration)

EC0 = 100 mg/L (Daphnia magna, 48 h) (nominal concentration)

EC100 > 100 mg/L (Daphnia magna, 24 h) (nominal concentration)

EC100 > 100 mg/L (Daphnia magna, 48 h) (nominal concentration)

Description of key information

Key value for chemical safety assessment

EC50/LC50 for freshwater invertebrates:
100 mg/L

Additional information