2,2,6,6-tetramethylpiperidinooxy

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2004-08-17 to 2004-09-20

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)

Version / remarks:

1999 (draft)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

1992

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Nominal test concentrations 100 mg/l; 50 mg/L; 25 mg/L, 12.5 mg/L; 6.25 mg/L; 3.13 mg/L; 1.56 mg/L
- Sampling method: Separate replicates for the test item analysis were prepared and incubated under test conditions. Concentrations were analytically verified at 0 h and 48 h by HPLC-analysis.
- Sample storage conditions before analysis: storage at 7 +/- 2°C until start of analysis if necessary.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Stock solution (100 mg/L) has been perpared with dilution water one day prior to application. The stock solution was stirred 24 h at room temperature
- Controls: dillution water without test item
- Evidence of undissolved material: The test item was clearly dissolved in the tested concentrations and was concentration related slightly yellowish coloured at the tested concentrations 50.0 and 100 mg/L throughout exposure.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna, water flea
- Strain/clone: Daphnia magna STRAUS (Clone 5)
- Source: lnstitute for Water, Soil, and Air hygiene (Germany), breed in the laboratory
- Culture: In 2-3 L glass vessels with approximatively 1.8 L culture medium, at 21 °C, in an incubator, 16 h illumination, illumination strength max. 20 µE·m-2·S-1.
- Culture medium: Culture medium Elendt M4, according to Elendt (1990).
- Feeding: At least 5x weekly ad libitum with a mix of Desmodesmus subspicatus and Chlorella vulgaris, with an algae cell density of> 10_6 cells/ml.
- Age of parental stock: At test start the animals were 2-24 h old and were no first brood progeny.
- Feeding during test: no feeding during study.

ACCLIMATION
- Acclimation period: Acclimation to the dilution water ca. 2 h before test start. 2 to 24 h old daphnids were used for the test. They were obtained by removing the mother animals twice within 22 h.
- Acclimation conditions: same as test
- Health during acclimation (any mortality observed): Daphnids were taken from a healthy stock.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

256 mg CaCO3/L

Test temperature:

20 °C, ± 1 °C constant

pH:

test start: 8.00-8.07, test end: 7.83-7.99

Dissolved oxygen:

7.45-8.10 mg O2/L; >5.21 mg/L

Nominal and measured concentrations:

Test Item: 1.56 - 3.13 - 6.25 - 12.5 - 25.0 - 50.0 - 100 mg/L
Reference Item: 0.58 - 1.0 - 1.8 - 3.2 - 5.8 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass beakers, 50 mL volume
- Material, size, headspace, fill volume: glass, 50 mL volume, 20 mL study medium
- Renewal rate of test solution: static
- No. of organisms per vessel: 5 organisms per vessel (10 organisms in the test with the reference item)
- No. of vessels per concentration (replicates): 4 (2 in the test with the reference item)
- No. of vessels per control (replicates): 4
- Biomass loading rate:

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: according to EC 92/69 L383A C.2. Annex
- Total organic carbon: 1.44 mg/L according to water analysis
- Culture medium different from test medium: Culture medium Elendt M4 (Elendt (1990)), modified to a total hardness of 160 to 180 mg CaC03/L.
- Intervals of water quality measurement: Water quality parameters of pH-value and dissolved oxygen concentration were measured at 0 h and 48 h of the test.

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16/8 h light/dark cycle
- Light intensity: max. 20 μE·m-2· s-1 (1200 lx)

EFFECT PARAMETERS MEASURED : immobilisation after 24 h and 48 h. Water parameters were measured at 0 h and 48 h


RANGE-FINDING STUDY
- Test concentrations: 0.1 to 100 mg test item/L (non-GLP)
- Results used to determine the conditions for the definitive study: Based on the results of the preliminary test the definitive test was performed with 7 nominal concentrations ranging from 1.56 to 100 mg/L in a geometric series with a factor of 2 under static conditions to enable the determination of immobilisation after 24 and 48 hours.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate p.a.

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

51.3 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks on result:

other: confidence interval (95%): 37.0 - 71.2 mg/L

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

27.4 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: confidence interval (95%): 24.2 - 31.1 mg/L

Duration:

24 h

Dose descriptor:

EC10

Effect conc.:

15.9 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

EC10

Effect conc.:

16.3 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

Duration:

48 h

Dose descriptor:

EC100

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

12.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Solubility of the test item: The test item was clearly dissolved in the tested concentrations and was concentration related slightly yellowish coloured at the tested concentrations 50.0 and 100 mg/L throughout exposure.
- Mortality of control: In the control group no Daphnia were immobilised or trapped on the surface of the water.

Water quality parameters of pH-value and dissolved oxygen concentration measured at 0 h and 48 h were determined to be within the acceptable limits. The dissolved 0 2 concentration was > 5.21 mg/L (60 % of the air saturation value at the temperature used). The pH-value did not vary by more than 1 unit.

Recovery rates of the test item were > 80 %. Therefore, all effects are given based on nominal test item concentrations.

Results with reference substance (positive control):

A reference test was carried out with potassium dichromate as reference item. The EC50-value of the reference item at 1.94 mg/L after 24 h is within the prescribed concentration range of 1.0 to 2.5 mg/L according to AQS, DIN Guideline 38412 L 30.

Reported statistics and error estimates:

The EC10 and EC50-values were calculated by probit analysis according to WEBER (1986).

Validity criteria fulfilled:

yes

Remarks:

The validity criteria of the test guideline are fulfilled.

Conclusions:

The Acute lmmobilisation of the test item 2,2,6,6-Tetramethyl-1-piperidinyloxyl (TEMPO) to Daphnia magna was determined according to OECD Guideline 202 (1999, Draft) and EC Directive 92/69/EC Method C.2. The study was conducted with 7 concentrations ranging from 1.56 to 100 mg/L in a geometric series with a factor of 2 under static conditions over a duration of 48 hours. 20 test organisms were exposed to each test concentration and control. All concentration levels of 2,2,6,6-Tetramethyl-1-piperidinyloxyl (TEMPO) and the control were analytically verified via HPLC after 0 h and 48 h. Recovery rates of the test item were > 80 %. Therefore, all effects are given based on nominal test item concentrations. The EC50 after 48 h has been determined to be 27.4 mg/L.

Description of key information

In a static acute toxicity test with daphnia, an EC50 of 27.4 mg/L has been determined after 48 h.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

27.4 mg/L

Additional information

An acute toxicity test on daphnia has been performed with the test substance 2,2,6,6-Tetramethyl-1-piperidinyloxyl according to the guideline TG OECD 202 and EU method C.2. Seven test substance concentrations ranging from 1.56 mg/L to 100 mg/L (factor 2) has been tested. 20 daphnids per test substance concentration were exposed to the test substance under static conditions for 48 h. The concentration levels of 2,2,6,6 -Tetramethyl-1 -piperidinyloxyl and the control were analytically verified by HPLC analysis after 0 h and 48 h. The recovery rates of the test substance were >80%. Therefore, all effects are based on the nominal test substance concentration. The EC50 has been determined to be 27.4 mg/L after 48 h. All validity criteria of the test guideline have been fulfilled.