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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:
March 2003
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Carried out using OECD guideline and under GLP (check certificate of analysis
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Qualifier:
according to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
A sample of 5 mL was taken from the LS solution container at the beginning of the test and stored at -20°C until analysis. Samples totaling 5 mL were also taken at T24 and T48 hours from the LS test vessels containing daphnids and stored at -20°C until analysis.
Vehicle:
no
Details on test solutions:
The stock solution, for the limit and range-finding tests, was prepared by dispersing the test item directly in M4 reconstituted water. 100 mg of test item was agitated for 23h in 1000ml of M4 to give a loading rate of 100 mg/L. As test item droplets were still in suspension after agitation, the stock solution was filtered through a filter of porosity 0.45 uM to remove the suspended fraction. The filtered solution, corresponding to the limit of solubility (LS) of the test item under the experimental conditions, was then used to prepare the test solutions.

Test solutions were prepared by further dilution of the LS solution with M4 reconstituted water to provide a geometric series of concentrations: 0 mg/L, LS/1000, LS/1 00, LS/10 and LS. Test vessels containing daphnids were filled directly from the test solution containers immediately after preparation and test solutions remained unchanged. The pH of the test solutions remained within acceptable limits (between 6.5 and 9 and in the range ± 0.5 unit of the test water) after preparation and there was no adjustment of pH before addition ofthe animals.
Test organisms (species):
Daphnia magna
Details on test organisms:
Species: Daphnia magna STRAUS-clone 5.
Reason for this choice: species commonly used in Europe for aquatic toxicity testing and recommended in OECD and EEC guidelines.
Breeder: CIT.
Origin: Centre Technique du Bois et de l'Ameublement, 75012 Paris, France.
Life stage: at the beginning of the test, first instar dapbnids between 6 and 24 hours old and produced from parthenogenically reproducing brood female population.
Acclimation: animals are held in water of the same quality as the test water from birth until use in the test.

Environmental conditions during culture
During the culture period, the stock aquarium conditions werer as follows:
Water: reconstituted (M4). The pH ofthis solution was 8.0 ± 0.5; the sum ofCa and Mg ions in the solution was 2.5 mmol/L (molar ratio Ca:Mg is approximately 4:1 and Na:K approximately 10:1); total hardness was generally between 238 and 289 mg/L as CaC03.

Food
Culture animals were fed with a diet of between 0.1 and 0.2 mg of carbon per Daphnia per day, in the form ofthe algal strain Scenedesmus subspicatus which is cultured at CIT. Animals were not fed during the test.

Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
48 h
Hardness:
238 - 289 mg/L as CaCO3
Test temperature:
18 - 22 oC
pH:
6.5 - 9
Dissolved oxygen:
>60%
Salinity:
not relevant for freshwater tests
Nominal and measured concentrations:
Loading rate = 100 mg/L
Details on test conditions:
Administration ofthe test item
The study included a limit and a range-finding tests.

Limit test
Twenty animals (in four replicates of five) were subjected to the LS solution (limit of solubility) obtained from an initial loading rate of approximately 100 mgiL) and a further group of 20 animals (in four replicates of five) was exposed to dilution water only (control), over a 48-hour period

Range-finding test
Three groups of 20 daphnids (in four replicates of five) were subjected to dilutions of the limit test solution at LS/1 000, LS/I00 and LS/1 0 for a period of 48 hours.

pH and dissolved oxygen
The pH and dissolved oxygen values of the control and the highest concentration were measured at the beginning and the end ofthe test. Temperature was measured at the beginning of the test and the maximum and minimum temperatures during the test recorded. Water hardness was checked at the beginning and the end of the test in the control solution.

Other test conditions
Photoperiod: 16 hrs light, 8 hrs dark
Reference substance (positive control):
no
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: based on initial loading rate of 100 mg/L
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
> 0.802 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Results with reference substance (positive control):
not applicable

Number of immobilized Daphnids

Immobilisation (n animals out of 20) at each nominal concentration *

 

0

LS/1000

LS100

LS/10

LS

 0 hr

0

0

0

0

0

24 hr

0

0

0

0

0

48 hr

0

0

0

0

0

Validity criteria fulfilled:
yes
Conclusions:
Under the experimental conditions, the 48-hour EC50 of 2,5-DIMETHYL-2,5-DI(2-ETHYLHEXANOYL PEROXy) HEXANE in a static test system is > 100 mg/L expressed as initial loading rate, corresponding to 0.802 mg/L expressed as actual concentration (geometric mean of measured concentrations during the test in the saturated solution obtained from this loading rate of 100 mg/L) for Daphnia magna.
Executive summary:

The acute toxicity of the test item 2,5-DlMETHYL-2,5-DI(2-ETHYLHEXANOYL PEROXY)HEXANE was evaluated in the Cladoceran, Daphnia magna STRAUS (clone 5) using a 48-hour static test according to OECD guideline (No. 202, 4th Apri11984) and Commission Directive 92/69IEEC (C.2,31 st July 1992). The criterion measured is the EC50 (Median Effective Concentration), a statistically derived concentration which is expected to cause immobility in 50% of the animals.

Methods

The test item was dispersed in reconstituted water (M4) prepared from deionized water with a conductivity <10 uS/cm. The total hardness of the reconstituted water was generally between 238 and 289 mg/L (as CaC03) and the pH is 8.0 ± 0.5.

Limit test

Twenty daphnids (in four replicates of five) were exposed to a nominal concentration of LS (the limit test concentration) for 48 hours while a second group of 20 daphnids (in four replicates of five) was exposed to test dilution water (M4) only (control solution).

Range finding test

Three groups of 20 daphnids (in four replicates of five) were subjected to dilutions of the limit test solution at LS/1000, LS/100 and LS/10 for a period of 48 hours.

Results

No immobilization was observed at immobilization was observed at 0 mg/L, LS/1 000, LS/100, LS/1 0 and LS throughout the test.

Conclusions

Under the experimental conditions, the 48-hour EC50 of 2,5-DIMETHYL-2,5-' DI(2-ETHYLHEXANOYL PEROXy) HEXANE in a static test system is>100 mg/L expressed as initial loading rate, corresponding to 0.802 mg/L expressed as actual concentration (geometric mean of measured concentrations during the test in the saturated solution obtained from this loading rate of 100 mg/L) for Daphnia magna.

Description of key information

One OECD 202 GLP study is available where the 48h EC50 > 100 mg/L (loading rate). This is above the water solubility of the substance.

Key value for chemical safety assessment

Additional information

The acute toxicity of the test item was evaluated in the Cladoceran, Daphnia magna STRAUS (clone 5) using a 48-hour static test according to OECD guideline (No. 202, 4th Apri11984) and Commission Directive92/69IEEC (C.2,31 st July 1992). Thecriterion measured is the EC50 (Median Effective Concentration), a statistically derived concentration which is expected to cause immobility in 50% of the animals.

The test item was dispersed in reconstituted water (M4) prepared from deionized water with a conductivity<10 uS/cm. The total hardness of the reconstituted water was generally between 238 and 289 mg/L (as CaC03) and the pH is 8.0±0.5. Twenty daphnids (in four replicates of five) were exposed to a nominal concentration of LS (the limit test concentration) for 48 hours while a second group of 20 daphnids (in four replicates of five) was exposed to test dilution water (M4) only (control solution).

Range finding test

Three groups of 20 daphnids (in four replicates of five) were subjected to dilutions of the limit test solution at LS/1000, LS/100 and LS/10 for a period of 48 hours.

Results

No immobilization was observed atimmobilization was observed at 0 mg/L,LS/1 000,LS/100,LS/1 0 and LS throughout the test.

Under the e xperimental conditions, the 48-hour EC50 of 2,5-DIMETHYL-2,5-' DI(2-ETHYLHEXANOYL PEROXy) HEXANE in a static test system is>100 mg/L expressed as initial loading rate, corresponding to 0.802 mg/L expressed as actual concentration (geometric mean of measured concentrations during the test in the saturated solution obtained from this loading rate of 100 mg/L) forDaphnia magna. In conclusion, no effect were seen to Daphnia magna in concentrations below the water solubility limit.