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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
Reliability:
1 (reliable without restriction)
Qualifier:
according to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
GLP compliance:
yes (incl. QA statement)
Analytical monitoring:
yes
Details on sampling:
A sample of each test concentration was taken for chemical analysis at 0 and 48 hours in order to determine the stability of the test item under test conditions. All samples were stored frozen prior to analysis. Only concentrations within the range to be used for the definitive test were analyzed.
Vehicle:
no
Details on test solutions:
Reconstituted water (ISO medium) was used for the range-finding.. Elendt M7 media, as used for the culturing, was used for the definitive test.

Reconstituted Water – Elendt M7 Medium

Ingredient
Final Concentration (mg/L)
H3BO3
0.715
MnCl2.4H2O
0.090
LiCl
0.077
RbCl
0.018
SrCl2.6H2O
0.038
NaBr
0.004
Na2MoO4.2H2O
0.016
CuCl2.2H2O
0.004
ZnCl2
0.013
CoCl2.6H2O
0.010
KI
0.0033
Na2SeO3
0.0022
NH4VO3
0.00058
Na2EDTA.2H2O
0.625
FeSO4.7H2O
0.249
CaCl2.2H2O
293.8
NaHCO3
64.8
MgSO4.7H2O
123.3
Na2SiO3.9H2O
10
KCl
5.8
NaNO3
0.274
K2HPO4
0.184
KH2PO4
0.143
Thiamine hydrochloride
0.075
Cyanocobalamine (vitamin B12)
0.0010
D(+) biotin (vitamin H)
0.00075
The pH of the prepared media was 7.9 ± 0.3 and stored at approximately 21 ºC.

Elendt M7 media, as used for the culturing, was used for the definitive test

Reconstituted Water – ISO Medium
Ingredient
Final Concentration (mg/L)
CaCl2.2H2O
294
MgSO4.7H2O
123
NaHCO3
65
KCl
5.8

Test organisms (species):
Daphnia magna
Details on test organisms:
The test was carried out using 1st instar Daphnia magna derived from in-house laboratory cultures.
Adult daphnia were maintained in 150 mL glass beakers containing Elendt M7 medium (see Annex 1) 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
Hardness:
See section with details on test solution
Test temperature:
18-22 C
pH:
See section with details on test solution
Dissolved oxygen:
See section with details on test solution
Salinity:
See section with details on test solution
Conductivity:
See section with details on test solution
Details on test conditions:
Range-finding Test
The results obtained from the preliminary media preparation trial conducted indicated that a dissolved test item concentration of approximately 90 mg/L could be obtained using a saturated solution method of preparation.
The test concentrations to be used in the definitive test were determined by a preliminary range-finding test.
In the range-finding test Daphnia magna were exposed to a series of nominal test concentrations of 0.10, 1.0, 10 and 100% v/v saturated solution.
A nominal amount of test item (1100 mg) was dispersed in 11 liters of test water with the aid of propeller stirring at approximately 1500 rpm for 24 hours. After 24 hours the stirring was stopped and any undissolved test item was removed by filtration through a 0.2 μm Sartorius Sartopore filter (first approximate 2 liters discarded in order to pre-condition the filter) to give a 100% v/v saturated solution. A series of dilutions was made from this saturated solution to give further test concentrations of 0.10, 10 and 10% v/v saturated solution.
Each prepared concentration was inverted several times to ensure adequate mixing and homogeneity.
In the range-finding test 10 daphnids were placed in each test and control vessel 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 and 8 hours darkness for a period of 48 hours with 20 minute dawn and dusk transition periods. 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.
The control group was maintained under identical conditions but not exposed to the test item.
A sample of each test concentration was taken for chemical analysis at 0 and 48 hours in order to determine the stability of the test item under test conditions. All samples were stored frozen prior to analysis. Only concentrations within the range to be used for the definitive test were analyzed.

Definitive Test
Based on the results of the range-finding test the following test concentrations were assigned to the definitive test: 2.6, 6.4, 16, 40 and 100% v/v saturated solution.
A nominal amount of test item (1100 mg) was dispersed in 11 liters of test water with the aid of propeller stirring at approximately 1500 rpm for 24 hours. After 24 hours the stirring was stopped and any undissolved test item was removed by filtration through a 0.2 μm Sartorius Sartopore filter (first approximate 2 liters discarded in order to pre-condition the filter) to give a 100% v/v saturated solution. A series of dilutions was made from this saturated solution to give further test concentrations of 2.6, 6.4, 16 and 40% v/v saturated solution.
Each prepared concentration was inverted several times to ensure adequate mixing and homogeneity.
The concentration and stability of the test item in the test preparations was verified by chemical analysis at 0 and 48 hours.
Exposure Conditions
As in the range-finding test 150 mL glass beakers containing approximately 100 mL of test preparation were used. At the start of the test 5 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.
The test preparations were not renewed during the exposure period.

The following deviation from study plan occurred:
The study plan states that the water temperature will be maintained at 18 to 22 °C with a maximum deviation of ±1 °C, however, at initiation of the definitive test the water temperatures in the 6.4, 16, 40 and 100% v/v saturated solution test groups were higher than 22 °C, with a maximum recorded temperature of 22.9 °C. This deviation was considered to have no impact on the outcome or integrity of the study.
Reference substance (positive control):
yes
Remarks:
A positive control (Envigo Study Number MS29MC) used potassium dichromate as the reference item. The positive control was conducted between 29 February 2016 and 02 March 2016.
Duration:
24 h
Dose descriptor:
EC50
Effect conc.:
> 20 - < 32 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 11 - < 17 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
24 h
Dose descriptor:
NOEC
Effect conc.:
6.4 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
6.4 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
24 h
Dose descriptor:
LOEC
Effect conc.:
16 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
LOEC
Effect conc.:
16 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
No immobilization was observed at the 0.10% v/v saturated solution test concentration, however, immobilization was observed at test concentrations of 1.0, 10 and 100% v/v saturated solution.
Based on this information test concentrations of 2.6, 6.4, 16, 40 and 100% v/v saturated solution were selected for the definitive test.
Chemical analysis of the 1.0, 10 and 100% v/v saturated solutions at 0 hours (see Annex 5) showed measured test concentrations to range from 104% to 137% of nominal concentrations. There was no significant change in the measured concentrations at 48 hours indicating that the test item was stable under test conditions.
Definitive Test
Verification of Test Concentrations
Chemical analysis of the test preparations at 0 hours showed measured test concentrations were within 80% to 120% of the nominal concentrations (ranging from 2.2 to 105 mg/L). There was no significant change in the measured concentrations at 48 hours and so the results are based on the nominal test concentrations only.
Immobilization Data
Analysis of the immobilization data by Probit analysis using Linear Maximum-Likelihood regression at 24 hours and by Trimmed Spearman-Karber procedure at 48 hours based on the nominal test concentrations gave the results presented in section "Effect concentration".
Results with reference substance (positive control):
A positive control (Envigo Study Number MS29MC) used potassium dichromate as the reference item at concentrations of 0.32, 0.56, 1.0, 1.8 and 3.2 mg/L.
Exposure conditions for the positive control were similar to those in the definitive test.
The results from the positive control with potassium dichromate were within the normal range for this reference item.
Validity criteria fulfilled:
yes
Conclusions:
Exposure of the freshwater invertebrate Daphnia magna to the 1,2-Ethanediamine, N-(2-aminoethyl)-, reaction products with glycidyl tolyl ether has been investigated and gave the following effect levels - EC50=25 mg/l (24 h exposure) and EC50=14 mg/l (084 h exposure), NOEL = 6.4 mg/l (24 h and 48 h exposure), LOEL=16 mg/l (24 h and 48 h exposure).
Executive summary:

Exposure of the freshwater invertebrate Daphnia magna to the 1,2-Ethanediamine, N-(2-aminoethyl)-, reaction products with glycidyl tolyl ether  has been investigated and gave the following effect levels  - EC50=25 mg/l (24 h exposure) and  EC50=14 mg/l (084 h exposure), NOEL = 6.4 mg/l  (24 h and 48 h exposure), LOEL=16 mg/l  (24 h and 48 h exposure).

Description of key information

Exposure of the freshwater invertebrate Daphnia magna to the 1,2-Ethanediamine, N-(2-aminoethyl)-, reaction products with glycidyl tolyl ether  has been investigated and gave the following effect levels  - EC50=25 mg/l (24 h exposure) and  EC50=14 mg/l (084 h exposure), NOEL = 6.4 mg/l  (24 h and 48 h exposure), LOEL=16 mg/l  (24 h and 48 h exposure).

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Effect concentration:
14 mg/L

Additional information