4-tert-Butoxystyrene

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:

2008

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:

April 13, 2004

Deviations:

yes

Remarks:

The spacing factor was enlarged. The study integrity was not adversely affected.

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 1.0, 3.2, 10, 32 and 100% of a water soluble fraction (WSF) prepared at a loading rate of 100 mg/l.
- Sampling method: singular samples for possible analysis were taken from all test concentrations and the control. Sampling vessels were pre-incubated with the respective test solutions before sampling. In addition, the glass wool used for preparation of the WSF was retained for possible analysis. At the end of the exposure period, the replicates were pooled at each concentration before sampling. Additionally, reserve samples of 2 ml were taken.
- Sample frequency and volume: at 0 h and 48 h; 2 ml from the approximate centre of the test vessels.
- Sample storage conditions before analysis: samples were stored in a freezer until analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method:
Butoxystyrene tested was not completely soluble in test medium at the loading rate prepared.
Preparation of test solutions started with a loading rate of 100 mg/l applying 24 hours of magnetic stirring in order to obtain maximum solubility of the test substance. Afterwards, the mixture was poured over glass wool to discard any undissolved material. The clear and colourless Water Soluble Fraction (WSF) was used for testing, together with several dilutions in ISO-medium.
One hour before the final test was started, a part of each test solution was used for pre-incubation of the respective test vessels.
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): the mixture was poured over glass wool to discard any undissolved material.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Age at study initiation: < 24 hours
- Stage and instar at study initiation: at least third generation
- Validity of batch: daphnids originated from a healthy stock, 2nd to 5th brood, showing no signs of stress such as mortality >20%, presence of males, ephippia or discoloured animals and there was no delay in the production of the first brood.
- Method of breeding: acyclical parthenogenesis.
- Source: in-house laboratory culture with a known history.
- Age of parental stock: more than 2 weeks old
- Feeding during test: no

METHOD FOR PREPARATION AND COLLECTION OF EARLY INSTARS OR OTHER LIFE STAGES, INCLUDING CULTURING CONDITIONS:
Each batch started with newborn daphnids (less than 3 days old) by placing about 250 of them into 5 litres of M7 medium in an all-glass culture vessel. The cultures reached a maximum age of 4 weeks and half of the medium was renewed twice a week after 7 days of cultivation. The temperature of the medium ranged from 18-22 °C. Feeding occurred daily and consisted of a suspension of fresh water algae.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

180 mg/l expressed as CaCO3

Test temperature:

19.9-20.3 °C. The temperature was continuously measured in a temperature control vessel.

pH:

7.7-8.0

Dissolved oxygen:

9.3-9.6 mg/l

Nominal and measured concentrations:

The average exposure concentrations were calculated to be 0.36, 1.1, 3.3, 11 and 36 mg/l at 1.0, 3.2, 10, 32 and 100% of the WSF prepared at a loading rate of 100 mg/l.

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 ml, all-glass
- Volume of test solution: 80 ml
- Aeration: no aeration of the test solutions
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: adjusted ISO medium
- M7 medium, as prescribed by Dr. Elendt-Schneider (Elendt, B.-P., 1990: Selenium deficiency in Crustacea. An ultrastructural approach to antennal damage in Daphnia magna Straus. Protoplasma 154, 25-33).

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 hours daily
- Introduction of daphnids: within 15 minutes after preparations of the test solutions.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Immobility: at 24h and at 48h.
- pH: at the beginning and at the end of the test for all concentrations and the control.
- Dissolved oxygen: at the beginning and at the end of the test for all concentrations and the control.

COMBINED LIMIT/RANGE-FINDING STUDY
- Test concentrations: 0.1, 1.0, 10 and 100% of the WSF prepared at a loading rate of 100 mg/l.
- Results used to determine the conditions for the definitive study: the expected LC50 was between 0.27 and 3.0 mg/l.

Reference substance (positive control):

yes

Remarks:

Reference substance: potassium dichromate (K2Cr2O7). Date of last reference test: April 2008.

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

7.3 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat. (dissolved fraction)

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

1.1 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat. (dissolved fraction)

Basis for effect:

mobility

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

3.8 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat. (dissolved fraction)

Basis for effect:

mobility

Results with reference substance (positive control):

- Results with reference substance valid? Yes.
- 48h-EC50: 0.30 mg/l, which is just within the allowed range of 0.3 and 1.0 mg/l.

Reported statistics and error estimates:

95% confidence intervals for 24h-EC50 and 48h-EC50 are 6.3-9.1 and 3.1-5.2 respectively.

Validity criteria fulfilled:

yes

Conclusions:

The 48h-EC50 was 3.8 mg/l based on average exposure concentrations.
Under the conditions of the present study butoxystyrene did not induce acute immobilisation of Daphnia magna at 1.1 mg/l after 48 hours of exposure (NOEC).

Executive summary:

Acute toxicity of butoxystyrene was assessed in Daphnia magna. Based on the results of a preceding combined limit/range-finding test a final test was performed. Preparation of test solutions started with a loading rate of 100 mg/l applying 24 hours of magnetic stirring in order to obtain maximum solubility of the test substance. Afterwards, the mixture was poured over glass wool to discard any undissolved material. The clear and colourless Water Soluble Fraction (WSF) was used for testing, together with several dilutions in ISO-medium. One hour before the test was started, a part of each test solution was used for pre-incubation of the respective test vessels.

Twenty daphnids per concentration (four replicates with five daphnids each) were exposed to a control group and to 1.0, 3.2, 10, 32 and 100% of the WSF prepared at a loading rate of 100 mg/l. The total test period was 48 hours and samples for analytical confirmation of actual test concentrations were taken at the start and the end of the test. The average exposure concentrations were calculated to be 0.36, 1.1, 3.3, 11, 36 mg/l.

The study met the acceptability criteria prescribed by the protocol and was considered valid. Butoxystyrene did not induce acute immobilisation of Daphnia magna at 1.1 mg/l after 48 hours of exposure (NOEC). The 48h-EC₅₀ was 3.8 mg/l based on average exposure concentrations.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Dose descriptor:

EC50

Remarks:

48h

Effect concentration:

3.8 mg/L

Additional information

The acute toxicity of butoxystyrene in Daphnia magna was assessed in a GLP-compliant study according to OECD 202. The study was performed under static conditions and the total test period was 48 hours. Based on the results of a preceding combined limit/range-finding test a final test was performed.  Twenty daphnids per concentration were exposed to a control group and to 1.0, 3.2, 10, 32 and 100% of a water soluble fraction (WSF) prepared at a loading rate of 100 mg/l. The average exposure concentrations were calculated to be 0.36, 1.1, 3.3, 11, 36 mg/l. No acute immobilisation of Daphnia magna was induced by butoxystyrene at 1.1 mg/l after 48h of exposure (NOEC). The 48h-EC₅₀ was 3.8 mg/l based on average exposure concentrations.