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EC number: 240-539-0 | CAS number: 16484-77-8
Short-term toxicity to aquatic invertebrates
Administrative data
Link to relevant study record(s)
- Endpoint:
- short-term toxicity to aquatic invertebrates
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 07 November 1990 to 09 November 1990
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- EU Method C.2 (Acute Toxicity for Daphnia)
- Deviations:
- no
- Qualifier:
- equivalent or similar to guideline
- Guideline:
- OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
- Version / remarks:
- April 4, 1984
- Deviations:
- no
- GLP compliance:
- yes
- Analytical monitoring:
- yes
- Details on sampling:
- - Concentrations: 12.5 and 100 mg/L. Also 0 mg/L for solvent control and control.
- Sampling method: At the start of the test (after 0 hours) samples from vessels which were not stocked with daphnids were anylsed and at the end of the test (after 48 h) samples from stocked and unstocked vessels were analysed.
- Sample storage conditions before analysis: Refrigerator - Vehicle:
- not specified
- Details on test solutions:
- PREPARATION AND APPLICATION OF TEST SOLUTION
The synthetic medium M4 (4) is used for the culture and test. The medium is prepared on the basis of an ultrapure, deionized water (conductivity < 0.05 µS/cm).
At the beginning of the test the stock solution of the test material was freshly prepared. A defined amount of the test material was weighed out and suspended directly in the test medium by stirring (5 min at 20 °C).
- Concentration of vehicle in test medium: The nominal concentration of the stock solution was 100 mg/L. By dilution of this stock solution with test medium the following nominal concentrations were prepared: 100 mg/L, 50 mg/L, 25 mg/L and 12.5 mg/L.
- Test organisms (species):
- Daphnia magna
- Details on test organisms:
- TEST ORGANISM
- Common name: Daphnia magna
- Strain/clone: STRAUS
- Age at study initiation: 2 - 24 h
- Age of stock aniamls: 2 - 4 weeks
- Method of breeding: The daphnids are cultured under standard conditions in the laboratory.
- Feeding during test : None
- Test type:
- static
- Water media type:
- freshwater
- Limit test:
- no
- Total exposure duration:
- 48 h
- Post exposure observation period:
- None
- Hardness:
- 2.20 - 3.20 (mmol/L)
- Test temperature:
- 20.0 ± 1.0 ºC
- pH:
- 7.5 - 8.5
- Dissolved oxygen:
- 7.86 - 9.44 mg/L
- Conductivity:
- 600 - 700 µS/cm
- Nominal and measured concentrations:
- Nominal: 100 mg/L, 50 mg/L, 25 mg/L and 12.5mg/L.
- Details on test conditions:
- TEST SYSTEM
- Test vessel: Test tubes (glass) with flat bottom. Nominal volume 20 mL
- Volume of solution: 10 mL
- Aeration: The medium was aerated until saturated with oxygen, and was left to stand for 24 hours for stabilisation.
- No. of organisms per vessel: 5
- No. of vessels per concentration: 4
- No. of vessels per control: 4
- Biomass loading rate: 1/2
TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The synthetic medium M4 (4) is used for the culture and test. The medium is prepared on the basis of an ultrapure, deionised water.
- Alkalinity: Up to pH 4.3: 0.90 ± 0.10 (mmol/L) and 0.80 - 1.00 (mmol/L)
- Conductivity: 600 - 700 µS/cm
- Macro Nutrients: 80.1 mg/L Ca; 12.2 mg/L Mg; 19.8 mg/L Na; 3.2 mg/L K; 1.0 mg/L Si; 144.9 mg/L Cl; 0.2 mg/L NO3; 0.2 mg/L PO4; 48.4 mg/L SO4; 47.1 mg/L HCO3.
- Vitamins: 75.0 µg/L Thiamine; 1.0 µg/L B12; 0.75 µg/L Biotin.
- Trace Nutrients: 0.5000 mg/L B; 0.2000 mg/L Fe; 0.1000 mg/L Mn; 0.0500 mg/L Li; 0.0500 mg/L Rb; 0.0500 mg/L Sr; 0.0250 mg/L Mo; 0.0125 mg/L Br; 0.0063 mg/L Cu; 0.0063 mg/L Zn; 0.0025 mg/L Co; 0.0025 mg/L J; 0.0020 mg/L Se; 0.0003 mg/L V; 2.5000 mg/L EDTA*2H2O
- Intervals of water quality measurement:
Measurement of temperature: After 0, 24, 48 h in an extra vessel close to the test vessels.
Measurement of pH and oxygen: After 0 and 48 h in one parallel at each concentration.
OTHER TEST CONDITIONS
- Photoperiod: Artificial light; day: night-rhythm 16: 8 hours
- Light intensity: About 5 - 6 µE/(m E+2 x s) in the range of 400 - 700 nm
EFFECT PARAMETERS MEASURED: Swimming ability of the test animals checked visually after 0, 3, 6, 24 and 48 h.
- Reference substance (positive control):
- not required
- Key result
- Duration:
- 48 h
- Dose descriptor:
- EC0
- Effect conc.:
- 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- mobility
- 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
- Key result
- Duration:
- 48 h
- Dose descriptor:
- EC100
- Effect conc.:
- > 100 mg/L
- Nominal / measured:
- nominal
- Conc. based on:
- test mat.
- Basis for effect:
- mobility
- Details on results:
- Instead of mortality the test criterium of the acute toxicity is immobilisation of the daphnids after 48 hours of exposure which is expressed as EC (effect concentration). Those animals were considered to be immobile which were unable to swim within 15 seconds after gentle agitation of the test vessel.
The following values were determined:
EC0 (48 h): 100 mg/L
EC50 (48 h): > 100 mg/L
EC100 (48 h): > 100 mg/L
In the control immobilisation was ≤10 %. Thus, the validity criteria were fulfilled by the test. - Results with reference substance (positive control):
- No positive control used.
- Reported statistics and error estimates:
- A statistical evaluation of the data was unneccesary, because immobilisation was ≤ 10 % at the highest concentration (100 mg/L) tested.
- Validity criteria fulfilled:
- yes
- Conclusions:
- Under the conditions of the study the 48 h EC50 in Daphnia was > 100 mg/L. The 48 h EC0 was 100 mg/L and the EC100 was > 100 mg/L.
- Executive summary:
The acute toxicity of the test material to Daphnia magna was assessed according to EU Method C.2. and in compliance with GLP in a 48 h static test.
The test substance was tested in the following range of concentrations: 12.5 to 100 mg/L. The dilution factor was 2.
Instead of mortality the test criterium of the acute toxicity is immobilisation of the daphnids after 48 hours of exposure which is expressed as EC (effect concentration). Those animals were considered to be immobile which were unable to swim within 15 seconds after gentle agitation of the test vessel.
The validity criteria were fulfilled by the test.
Under the conditions of the study the 48 h EC50 in Daphnia was > 100 mg/L. The 48 h EC0 was 100 mg/L and the EC100 was > 100 mg/L.
- Endpoint:
- short-term toxicity to aquatic invertebrates
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 04 May 1994 to 06 May 1994
- 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)
- Deviations:
- not specified
- Qualifier:
- according to guideline
- Guideline:
- EU Method C.2 (Acute Toxicity for Daphnia)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Specific details on test material used for the study:
- Purity: 89.7 %
- Analytical monitoring:
- yes
- Details on sampling:
- Test concentrations were verified by chemical analysis. Water samples (200 mL in duplicate) were taken from the control and each exposure level (replicates pooled) at 0 hours and 48 hours and sent to the HRC Department of Environmental Analysis.
- Vehicle:
- no
- Details on test solutions:
- PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: The test material was dispersed in test water and stirred for 2 hours to give an initial stock solution of 100 mg/L. This solution was passed through a sieve (of approximately 0.5 mm^2 mesh size) to remove particulate matter before serial dilutions were prepared with test water to give the desired series of exposure levels. - Test organisms (species):
- Daphnia magna
- Details on test organisms:
- TEST ORGANISM
- Strain/clone: Straus.
- Age at study initiation: < 24 h
- Stage and instar at study initiation: Ist instar.
- Method of breeding: Parthenogenesis
- Source: Laboratory culture originating from a strain supplied by the Institute National de Recherche Chimique Appliquee (IRChA), France.
METHOD FOR PREPARATION AND COLLECTION OF EARLY INSTARS OR OTHER LIFE STAGES, INCLUDING CULTURING CONDITIONS: Brood stocks of Daphnia magna were cultured under a 16 h light: 8 h dark photoperiod at 23 ± 2 ° C in polypropylene vessels containing two litres of Elendt M7 medium. Cultures were fed daily with a suspension of mixed algae (predominantly Scenedesmus and Selenastrum spp.). Culture conditions ensure that reproduction is by parthenogenesis. Gravid adults were isolated 24 hours prior to initiation of the test. Young daphnids produced overnight were used for testing. - Test type:
- static
- Water media type:
- freshwater
- Limit test:
- no
- Total exposure duration:
- 48 h
- Test temperature:
- 22 °C
- pH:
- 7.0 - 7.8
- Dissolved oxygen:
- 8.2 - 8.4 mg/L
- Nominal and measured concentrations:
- Nominal: 0, 1, 2.2, 4.6, 10, 22, 46 and 100 mg/L test material (as supplied).
Calculated test material concentrations, adjusted for purity: 0.83, 2.0, 3.9, 9.0, 20, 42 and 91 mg/L. - Details on test conditions:
- TEST SYSTEM
- Test vessel: Jar
- Type: The jars were loosely covered with aluminium foil to minimise evaporation losses.
- Material, size, headspace, fill volume: Glass jar containing 200 mL of prepared test medium or diluent water only, as appropriate.
- Volume of solution: 20 mL test solution per organism.
- Aeration: Without supplementary aeration.
- Renewal rate of test solution (frequency/flow rate): Daphnia were exposed to the test or control conditions for a period of 48 hours without renewal of test media.
- No. of organisms per vessel: 20 animals per test group
- No. of vessels per concentration: Duplicate
- No. of vessels per control: Duplicate
TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt M7 medium was prepared using analytical grade reagents and reverse osmosis purified water.
Trace elements: 0.71 mg/L H3BO3, 0.090 mg/L MnCl2.4H2O, 0.077 mg/L LiCl, 0.018 mg/L RbCl, 0.038 mg/L SrCl2.6H2O, 0.0040 mg/L NaBr, 0.016 mg/L Na2MoO4.2H2O, 0.0042 mg/L CuCl.2H2O, 0.013 mg/L ZnCl2, 0.010 mg/L CoCl2.6H2O, 0.0033 mg/L Kl, 0.0022 mg/L Na2SeO3, 0.00058 mg/L NH4VO3, 1.7 mg/L Fe-EDTA solution.
Macro nutrients: 294 mg/L CaCl2.2H2O, 123 mg/L MgSO4.7H2O, 5.80 mg/L KCl, 64.8 mg/L NaHCO3, 6.83 mg/L Na2SiO3.5H2O, 0.274 mg/L NaNO3, 0.143 mg/L KH3PO4, 0.184 mg/L K2HPO4.
Vitamins: 0.075 mg/L thiamine hydrochloride, 0.0010 mg/L cyanocohalamine (B 12), 0.00075 mg/L biotin.
The above analytical grade reagents are dissolved in reverse osmosis purified water.
- Intervals of water quality measurement: The temperature in each vessel was measured daily and the pH and dissolved oxygen levels recorded at the start and at the end of the study.
OTHER TEST CONDITIONS
- Photoperiod: Photoperiod of 16 hours light: 8 hours dark.
EFFECT PARAMETERS MEASURED
- Daphnia were considered to be immobilised if they were unable to swim for approximately 15 seconds after gentle agitation. - Reference substance (positive control):
- no
- Duration:
- 48 h
- Dose descriptor:
- EC50
- Effect conc.:
- > 91 mg/L
- Nominal / measured:
- meas. (not specified)
- Conc. based on:
- test mat.
- Basis for effect:
- mortality
- Duration:
- 48 h
- Dose descriptor:
- NOEC
- Effect conc.:
- 91 mg/L
- Nominal / measured:
- meas. (not specified)
- Conc. based on:
- test mat.
- Basis for effect:
- mortality
- Details on results:
- Analysis of the immobility data gave the following results:
24 h EC50 > 91 mg/L
48 h EC50 > 91 mg/L
Lowest test concentration resulting in 100 % immobilisation: > 91 mg/L
The "no-observed effect level" was 48 h was 91 mg/L.
The statistical non-significance of the numbers of immobilised daphnids at the 2.2, 46 and 100 mg/L was verified using Fisher's Exact Test (P=0.5000, P=0.1154, P=0.5000 respectively).
Individual pH, temperature and dissolved oxygen values remained within acceptable limits throughout the duration of the study. - Reported statistics and error estimates:
- EC50 values and 95 % confidence limits are calculated according to the method of Thompson and Weil (1952). The "no-effect level" is determined by statistical analysis using Fisher's Exact Test (1973).
- Validity criteria fulfilled:
- not specified
- Remarks:
- expected to be valid source
- Conclusions:
- Under the conditions of the study the 48h-EC50 > 91 mg/L and a 48h-NOEC of 91 mg/L were determined on Daphnia magna.
- Executive summary:
The acute toxicity of the test was material to aquatic invertebrates was assessed according to EEC Methods for Determination of Ecotoxicity Annex to Directive 92/69/EEC (0.J No. L383A, 29 .12.92) Part C, Method 2 Acute toxicity for Daphnia and OECD Guideline for Testing of Chemicals No. 202, Part 1 Daphnia, Acute Immobilisation Test" and in compliance with GLP.
Groups of twenty, 1st instar Daphnia (less than 24 hours old) were exposed for 48 hours to seven concentrations of the test material dispersed in Elendt M7 medium. The incidence of immobilisation was recorded for each test and control group at 24 hours and at 48 hours and the following values determined:
24 h EC50 > 91 mg/L
48 h EC50 > 91 mg/L
Lowest test concentration resulting in 100 % immobilisation: > 91 mg/L
The "no-observed effect level was 48 h was > 91 mg/L
All results based upon measured concentrations expressed as the calculated equivalent concentration of the test material (as supplied). Measured concentrations ranged from 85 - 92 % of nominal at 0 hours and 81 - 90 % of nominal at 48 hours.
Under the conditions of the study the 48h-EC50 > 91 mg/L and a 48h-NOEC of 91 mg/L were determined on Daphnia magna.
Referenceopen allclose all
Summary of the Effective Concentrations of the Test Material Which Caused 0 %, 50 %, or 100 % Immobilisation.
Time (h) |
EC0 (mg/L) |
EC50 (mg/L) |
CL 95 % |
EC100 (mg/L) |
3 |
100 |
> 100 |
- |
> 100 |
6 |
100 |
> 100 |
- |
> 100 |
24 |
100 |
> 100 |
- |
> 100 |
28 |
100 |
> 100 |
- |
> 100 |
CL: Confidence limits
The EC values are given as nominal concentrations.
Minimum and Maximum Values of the Characteristics of the Test Solutions.
Parameter |
Minimum |
Maximum |
pH |
7.29 |
8.00 |
Oxygen content (mg/L) |
7.86 |
9.44 |
Temperature (°C) |
19.5 |
20.6 |
Summary of the Analysis of the Concentrations of the Test Material.
Time (h) |
Stocked with Daphnids |
Minimum (%) |
Maximum (%) |
0 |
No |
97.6 |
101.1 |
48 |
No |
102.5 |
103.5 |
48 |
Yes |
100 |
101.2 |
The values given above are the relative minimum and maximum concentrations of the test material in the test solutions at various times of the study. The values are given in percent of the nominal concentrations.
Cumulative Immobilisation Data forDaphnia magnaExposed for 48 Hours to the Test Material (As Supplied).
Nominal Concentration (mg/L) |
Cumulative Immobilised Daphnia (Initial Population: 10/Replicate) |
|||||||
24 h |
48 h |
|||||||
R1 |
R2 |
Total |
% |
R1 |
R2 |
Total |
% |
|
Control |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
1.0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
2.2 |
1 |
0 |
1 |
5 |
1 |
0 |
1 |
5 |
4.6 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
10 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
22 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
0 |
46 |
1 |
2 |
3 |
15 |
1 |
2 |
3 |
15 |
100 |
0 |
1 |
1 |
5 |
0 |
1 |
1 |
5 |
R1: Replicate 1
R2: Replicate 2
Environmental Parameters Mean Values for pH, Temperature and Dissolved Oxygen.
Nominal Concentration (mg/L) |
Mean Values |
||
pH |
mg O2/L |
T °C |
|
Control |
7.5 |
8.4 |
22 |
1.0 |
7.8 |
8.3 |
22 |
2.2 |
7.8 |
8.3 |
22 |
4.6 |
7.7 |
8.3 |
22 |
10 |
7.6 |
8.2 |
22 |
22 |
7.4 |
8.3 |
22 |
46 |
7.2 |
8.3 |
22 |
100 |
7.0 |
8.3 |
22 |
Measured Concentrations Mean Values and Percentages of Nominal.
Nominal Concentration Test Material (As Supplied) (mg/L) |
Number of Samples Analysed |
Calculated* Concentration of Test Material (mg/L) |
Concentration of Test Material (as Supplied) (% Nominal) |
Control |
2 |
- |
- |
1.0 |
2 |
0.8299 |
83 |
2.2 |
2 |
1.982 |
90 |
4.6 |
2 |
3.909 |
85 |
10 |
2 |
9.306 |
90 |
22 |
2 |
20.06 |
91 |
46 |
2 |
41.60 |
90 |
100 |
2 |
90.95 |
91 |
*Concentration adjusted for purity: Calculated concentration = measured concentration x 100 / 89.7
Description of key information
Jatzek (1993)
Under the conditions of the study the 48 h EC50 in Daphnia was > 100 mg/L. The 48 h EC0 was 100 mg/L and the EC100 was > 100 mg/L.
Bell (1994)
Under the conditions of the study the 48h-EC50 > 91 mg/L and a 48h-NOEC of 91 mg/L were determined on Daphnia magna.
Key value for chemical safety assessment
Fresh water invertebrates
Fresh water invertebrates
- Effect concentration:
- 91 mg/L
Additional information
Jatzek (1993)
The acute toxicity of the test material to Daphnia magna was assessed according to EU Method C.2. and in compliance with GLP in a 48 h static test. The study was awarded a reliability score of 1 in accordance with the criteria set forth by Klimisch et al. (1997).
The test substance was tested in the following range of concentrations: 12.5 to 100 mg/L. The dilution factor was 2.
Instead of mortality the test criterium of the acute toxicity is immobilisation of the daphnids after 48 hours of exposure which is expressed as EC (effect concentration). Those animals were considered to be immobile which were unable to swim within 15 seconds after gentle agitation of the test vessel.
The validity criteria were fulfilled by the test.
Under the conditions of the study the 48 h EC50 in Daphnia was > 100 mg/L. The 48 h EC0 was 100 mg/L and the EC100 was > 100 mg/L.
Bell (1994)
The acute toxicity of the test was material to aquatic invertebrates was assessed according to EEC Methods for Determination of Ecotoxicity Annex to Directive 92/69/EEC (0.J No. L383A, 29 .12.92) Part C, Method 2 Acute toxicity for Daphnia and OECD Guideline for Testing of Chemicals No. 202, Part 1 Daphnia, Acute Immobilisation Test" and in compliance with GLP. The study was awarded a reliability score of 1 in accordance with the criteria set forth by Klimisch et al. (1997).
Groups of twenty, 1st instar Daphnia (less than 24 hours old) were exposed for 48 hours to seven concentrations of the test material dispersed in Elendt M7 medium. The incidence of immobilisation was recorded for each test and control group at 24 hours and at 48 hours and the following values determined:
24 h EC50 > 91 mg/L
48 h EC50 > 91 mg/L
Lowest test concentration resulting in 100 % immobilisation: > 91 mg/L
The "no-observed effect level was 48 h was > 91 mg/L
All results based upon measured concentrations expressed as the calculated equivalent concentration of the test material (as supplied). Measured concentrations ranged from 85 - 92 % of nominal at 0 hours and 81 - 90 % of nominal at 48 hours.
Under the conditions of the study the 48h-EC50 > 91 mg/L and a 48h-NOEC of 91 mg/L were determined on Daphnia magna.
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