4-tert-butyl-3-hydroxy-2,6-xylylacetonitrile

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study in compliance with GLP-criteria.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

4-tert-butyl-3-hydroxy-2,6-xylylacetonitrile

EC Number:

259-766-1

EC Name:

4-tert-butyl-3-hydroxy-2,6-xylylacetonitrile

Cas Number:

55699-10-0

Molecular formula:

C14H19NO

IUPAC Name:

2-(4-tert-butyl-3-hydroxy-2,6-dimethylphenyl)acetonitrile

Details on test material:

- Name of test material (as cited in study report): 4-tert-Butyl-3-hydroxy-2,6-xylylacetonitrile
- Physical state/ Appearance: Solid / white to yellowish
- Homogeneity: Homogeneous
- Storage conditions: Storage at room temperature

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

The appearance of the test solutions and dissolution behaviour of the test substance was observed and recorded daily. At the start of the exposure (0 h) samples from vessels without daphnids and at the end of the exposure (48 h) samples from vessels with daphnids were analysed. The transport to the Analytical Laboratory was done on the day of sampling.

Test solutions

Vehicle:

yes

Details on test solutions:

For correct dosing of the test substance acetone was used as solvent. An acetonic solution of 250.1 mg test substance in 50 mL acetone was
prepared. The acetonic stock solution was distributed on the bottom of a 1 L beaker to maximize the surface and the acetone was completely evaporated. For the control, 7.4 mL of acetone only was added to a beaker and evaporated. After this 800 mL test medium were added and the beakers are shaken (at 50 rpm) for about one day at the test temperature. Undissolved test substance was removed by filtration with a membrane filter (pore width 0.2 μm) and then the test solution was distributed into the test vessels. The control test solution was similarly prepared and treated. All test solutions were visibly clear over the exposure period. No adjustment of the pH during the test.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM:
- Test species: Daphnia magna STRAUS
- Origin: The clone of Daphnia magna STRAUS 1820 used was supplied by the Institut National de Recherche Chimique Appliquée, France, in 1978.
From this date on this clone was cultured and bred continuously in the Laboratory of Ecotoxicology of the Experimental Toxicology and Ecology of BASF SE in Ludwigshafen.
- Age of the animals at the start of exposure: < 24 hours (at least 3rd brood progeny)
- Age of parent animals: 2 – 4 weeks

CULTURE CONDITIONS:
- Photoperiod: 16 hours light, 8 hours dark
- Culture media: synthetic fresh water (Elendt M4) prepared according to OECD 202.
- Temperature: 20 ± 1 °C
- Diet: unicellular algae Desmodesmus subspicatus, cultured continuously at the test facility
- Feeding: daily
- Renewal of the culture water: twice weekly

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Test conditions

Hardness:

2.20 - 3.20 mmol/L

Test temperature:

21.1 - 21.5 °C

pH:

8.1 - 8.2

Dissolved oxygen:

8.1 - 8.7 mg O2/L

Nominal and measured concentrations:

- Nominal concentrations: 0, 1.0, 2.2, 4.6, 10, 22 and 46 mg/L (based on a range finding test with an 48-h EC50 value between 1-100 mg/L).
- Measured concentrations: 0, 0.94, 1.5, 3.5, 5.1, 5.5 and 7.1 mg/L

Details on test conditions:

TEST SYSTEM:
- Test vessel: Glass beakers (nominal volume 100 mL)
- Number of replicates per concentration: 4 replicates with test animals (5 animals per replicate)
- Total number of animals/concentration: 20
- Number of animals/vessel: 5
- Test volume: 50 mL
- Loading (biological): 0.1 animals/mL (the daphnids were introduced into the test vessel impartially)

TEST MEDIUM / WATER PARAMETERS:
- A synthetic fresh water (Elendt M4) is used as media for culture and test purposes. This M4 medium is prepared as described in OECD 202. The general properties of this medium are as follows.
- Total hardness: 2.20 – 3.20 mmol/L
- Acid capacity up to pH 4.3: 0.80 – 1.00 mmol/L
- Molar ratio Ca:Mg: about 4 : 1
- pH value: 7.5 – 8.5
- Conductivity: 550 - 650 μS/cm
- Dissolved oxygen: Must remain ≥ 3mg/L during the test. To assure optimal dissolved oxygen levels, the M4 medium is aerated for approximately 24 hours prior to use.

OTHER TEST CONDITIONS:
- Feeding: None
- Light intensity / Photo period: About 20 – 850 lux at a wave length of 400 -750 nm;
16 hours light : 8 hours darkness
- Aeration None

OBSERVATIONS:
- Each test vessel was visually checked for immobilised daphnids after 0, 24 and 48 hours. In addition any abnormal behaviour or appearance was documented.
- The appearance of the test solutions and dissolution behaviour of the test substance was observed and recorded daily.
- The chemical and physical parameters of the test medium (total hardness, acid capacity, pH and conductivity) were determined after aeration and prior to use in the test.
- Measurement of the pH and oxygen content at the start of the exposure and at the end of the exposure in replicate 1 of all test concentrations and the control.
- Measurement of temperature continuously during the whole period in a separate vessel filled with water proximal to the test vessels.

Reference substance (positive control):

yes

Remarks:

potassiumdichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

3.86 mg/L

Nominal / measured:

meas. (not specified)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% C.L.: 3.35-4.23 mg/L

Effect concentrations 2

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

6.21 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

No additional adverse effects or abnormal behaviour were observed in any of the test treatments.

The analytically determined concentrations of the test substance in the test water were markedly lower than the nominal concentration most likely due to the poor solubility of the test substance in test media. However, all the measured concentrations were within 20% of the initial measured concentrations. Following recommendations in OECD 202 the results are expressed as mean measured concentrations.

Results with reference substance (positive control):

Reference substance testst are conducted monthly. The last test before the study was performed 29 Jul 2008. From this test a 24-h EC50 of 0.63 mg/L was determined. This result is within the range of 0.6 – 2.1 mg/L (ISO 6341) indicating that the culture of Daphnia magna used in this study is responding normally to toxic stress.

Reported statistics and error estimates:

For the statistical evaluation of the data and calculation of the EC50 the probit method based on Finney (1971) was used with SAS statistical software.

Any other information on results incl. tables

Number of mobile daphnids:

Nominal concentration (mg/L)	Total mobile after 24h	Total mobile after 48h
0 (control)	20	20
1.0	20	20
2.2	20	20
4.6	20	13
10	13	3
22	10	1
46	1	0