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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:
2012-2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study (OECD 202) without deviation
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
yes
Details on sampling:
Storage of samples: None
Samples treatment: Dilution 2 to 5 with acetonitrile
Vehicle:
no
Details on test solutions:
Stock solutions of TERT-BUTYL CUMYL PEROXIDE were prepared by adding the convenient amount of test item into dilution water, using a “slow stirring” flask in order to avoid any emulsion. After 24 hours of stirring the aqueous phases were collected and conveniently diluted.

For the range-finding test, 69.9 µL (65.5 mg) of test item were added to 655 mL of dilution water.
For the definitive test, 107 µL (100 mg) of test item were added to 1 L of dilution water.
Test organisms (species):
Daphnia magna
Details on test organisms:
Daphnia magna Straus (Cladocera, Crustacea), clone 5 or clone A. Animals come from a stock breading in the laboratory reared in VOLVIC® water added with vitamin B12, Na2SeO3, 5H2O, spiruline, Calcium, Magnesium unicellular green fresh-water algae Pseudokirchneriella subcapitata and Chlorella Vulgaris. Selection of test organisms less than 24 hours old is made by filtration
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
No data
Test temperature:
20.2 ± 0.25°C
pH:
see "any other information on results incl. tables"
Dissolved oxygen:
see "any other information on results incl. tables"
Salinity:
Not relevant
Nominal and measured concentrations:
Preliminary test: nominal concentrations 100, 50, 10, 5, 1 and 0.5 mg/L
Definitive test: nominal concentrations: 50, 38.3, 29.4, 22.5, 17.3, 13.2 and 10 mg/L.

Measured concentration: see "any other information on results incl. tables"
Details on test conditions:
The study was performed in a thermostated room using 30 mL glass bottles stoppered with PTFE bungs and sealed with aluminium caps. The temperature, continuously recorded in one test flask, remained at 20.2 ± 0.25°C.
Physical-chemical parameters were measured using a METTLER TOLEDO 345 pH-meter for measurement of pH and with a WTW OXI 538 oxymeter for dissolved oxygen measurement. Water: ultra-pure water was used for the preparation of dilution water (resistivity > 18M).

Preliminary test

Approximately 90% of the final volume of the test solution was added into test flasks. Five Daphnia magna aged from 6 to 24 hours were added into each test flask, which were then totally filled with the test solution (30 mL). Two replicates were prepared for each concentration. Two control flasks were prepared under the same conditions, with no test item. Test flasks were incubated in darkness at a temperature of 20.9 °C. After 48 hours of incubation, mobile Daphnia magna were counted.

Dissolved O2 and pH were measured at the highest tested concentration and in the control.

Definitive test

Volumetric flasks were filled with the required volumes of the test item stock solution and dilution water in order to obtain the test item solutions at concentrations varying from 50 mg/l to 10 mg/L. The definitive test was performed with a concentration range forming a geometric progression with a factor of 1.3. Approximately 90% of the final volume of the test solution was added into test flasks. Five Daphnia magna aged from 6 to 24 hours were added into each test flask, which were then totally filled with the test solution (30 mL). Four replicates were prepared for each concentration. Four control flasks were prepared under the same conditions, with no test item. Test flasks were incubated in darkness at a temperature of 20.2 ± 0.25°C.

Dissolved O2 and pH were measured at the highest concentration and in the control at the beginning and at all concentration and in the control the end of the test.
Reference substance (positive control):
yes
Remarks:
potassium dichromate
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
4.3 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
3.44 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
The appearance of the test solutions was visually checked at the beginning and at the end of the test. Solutions were found to be clear, colourless over the period of the test. No precipitation was observed at the end of the test.
Results with reference substance (positive control):
The sensitivity of the test system and the methodology are evaluated every month by performing an inhibition test on sodium dichromate. The nearest value of EC50 obtained on September 9, 2012 was 0.75 mg/L. For information, ISO 6341 has a validity criteria specifying that EC50 must be in the range 0.6 to 2.1 mg/L.
Reported statistics and error estimates:
The inhibition of mobility data are analyzed using an Excel program. It was designed to calculate the EC50 value and the 95% confidence interval. Probit analysis is generally used to calculate the 24 and 48-hour EC50 values. The No-Observed-Effect Concentration (NOEC) is determined by statistic analysis using the Dunnett’s test (US EPA).

Dissolved O2 and pH measured at the beginning and at the end (48h) of the definitive test

 

 

Nominal concentration mg/L

pH

Dissolved O2 (mg/L)

T0

T48h

T0

T48h

0 (T)

8.11

7.72

8.9

8.6

10

8.12

7.69

9.0

8.4

13.2

8.10

7.70

9.0

8.5

17.3

8.09

7.72

9.0

8.5

22.5

8.09

7.70

9.1

8.4

29.4

8.08

7.64

9.1

8.1

38.3

8.07

7.65

9.0

8.3

50

8.07

7.66

9.0

8.2

 

 

Nominal and measured concentrations of the test item at the beginning and at the end of the exposure period

 

Concentrations of TERT-BUTYL CUMYL PEROXIDE

Nominal

Concentration

 

(mg/L)

Measured in non inoculated solutions

Initial

(mg/L)

Final

(mg/L)

Final/Initial

%

0 (T)

< DL

< DL

-

10

0.65

0.67

103

13.2

1.01

1.08

107

17.3

1.38

1.55

112

22.5

1.84

2.11

115

29.4

2.55

2.93

114.9

38.3

3.44

3.88

112.8

50

4.96

5.21

105

 

< DL : concentration lower than the Detection Limit of the analytical method (0.03 mg/L).

< QL : concentration lower than the Quantification Limit of the analytical method (0.11 mg/L).

 

The final concentration of TERT-BUTYL CUMYL PEROXIDE was within the designated limit of 80 % of the initial concentration in non-inoculated flasks.Thus, the test item concentration was maintained throughout the duration of the test.

 

Since the final measured concentrations in non-inoculated flasks were found to be lower than 80 % of the nominal concentration, estimation of effective concentrations was based on initial measured concentrations.

Validity criteria fulfilled:
yes
Conclusions:
48h-EC50= 4.3 mg/L
Executive summary:

The acute toxicity (inhibition of mobility) of the test item TERT-BUTYL CUMYL PEROXIDE to aquatic invertebrates for a duration of 48 hours was assessed according to the OECD Guideline 202 and GLP requirements. Daphnia magna were exposed under static conditions.

 

The results were as follows (based on initial measured concentrations):

48h-EC50= 4.3 mg/L

Description of key information

The acute toxicity to aquatic invertebrates was assessed according to the OECD Guideline 202 and GLP requirements: 48h-EC50= 4.3 mg/L.

Key value for chemical safety assessment

EC50/LC50 for freshwater invertebrates:
4.3 mg/L

Additional information

The acute toxicity (inhibition of mobility) of the test item TERT-BUTYL CUMYL PEROXIDE to aquatic invertebrates for a duration of 48 hours was assessed according to the OECD Guideline 202 and GLP requirements. Daphnia magna were exposed under static conditions.

 

The results were as follows (based on initial measured concentrations):

48h-EC50= 4.3 mg/L