6,6'-di-tert-butyl-2,2'-methylenedi-p-cresol

Long-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1999-02-15 till 1999-06-30

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to other study

Qualifier:

according to guideline

Guideline:

other: OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test) (April, 1984)

Deviations:

no

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

Version / remarks:

(April, 1997)

Deviations:

no

Principles of method if other than guideline:

not applicable

GLP compliance:

yes

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material: not applicable

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0.10, 0.22, 0.46, 1.0 and 2.2 mg/L of 2,2'-methylenebis(6-tert-butyl-4-methylphenol) plus control and solvent control
- Sampling method: After three days, the test solution was collected and extracted with hexane analytically measured by GC-MS.
- Samples of newly prepared exposure water were taken at day 2, 9, 16 as well as exposure water before reneval at day 5, 12 and 19.

Vehicle:

yes

Details on test solutions:

2,2'-methylenebis(6-tert-butyl-4-methylphenol) was mixed with DMF/Castor oil HCO-40 (OECD SIDS 6,6'-DI-TERT-BUTYL-2,2'-METHYLENEDI-P-CRESOL)
UNEP Publications 93 (3/1(w/w)) and dissolved into dilution water, dechlorinated tap waterof the Takarazuka city, to prepare the nominal concentrations described above (No precipitations as well as emulsions were observed in all the exposure levels). To set the dispersant concentrations equal (i.e. 50 uL/L) between the nominal concentrations, DMF/Castor oil HCO-40 solution was also used for the preparations.

Test organisms (species):

Daphnia magna

Details on test organisms:

Obtained form National Institute for Environmental Studies, and subcultured at the testing facility.
A matured female daphnia was selected from the subculture. On the next day, newly hatched juveniles were transferred into other culture vessels and further cultured for more than two weeks under the following conditions; temperature 20 +/- 1 °C, photoperiod 16h light – 8h dark, culture density 20 individuals per one liter of dilution water, feeding 0.1 to 0.2 mg carbon per day with Chlorella vulgaris. Newly hatched juveniles from, thus cultured, parents (with 0 % mortality during this culture period) were used in the experiment.

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

21 d

Post exposure observation period:

not reported

Hardness:

ranged from 60 to 85 mg/L (as CaCO3) = 3.4 - 4.8 °dH

Test temperature:

20.0 to 20.1°C measured at each vessel during of the test.

pH:

8.0 to 8.3 measured at each vessel during of the test.

Dissolved oxygen:

7.6 to 8.7 mg/L (more than 60% of saturation) measured at each vessel during of the test.

Salinity:

not reported

Nominal and measured concentrations:

nominal concentrations: 0.10, 0.22, 0.46, 1.0, 2.2 mg/L, solvent control and control

Details on test conditions:

- Test system: Semi-static (Three renewals per week; on Mon., Wed. and Fri.)
- Dilution water: dechlorinated tap water (pH 8.1, hardness 55.2 mg/L as CaCO3 = 3.1 °dH).
- Test volume for exposure: 80 mL (in 100 mL beaker)
- Number of replicate, individuals per replicate: Ten replicates, One daphnia per replicate.
- Water chemistry during exposure: Measurements of temperature, pH, DO and total hardness were conducted at day 0, 9 and 16 for newly prepared exposure solution and day 2, 12 and 19 for exposure solution before renewal.
- Temperature: 20 +/- 1 °C
- Photoperiod: 16h light / 8h dark
- Analysis: GC-MS
- Test period: 21 days

Reference substance (positive control):

yes

Remarks:

Potassium dichromate, Lot No. ACQ2610, reagent grade from Wako Pure Chemical Industries

Duration:

21 d

Dose descriptor:

LC50

Effect conc.:

1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Remarks on result:

other: 95% confidence limit: 0.71 - 1.4

Duration:

21 d

Dose descriptor:

EC50

Effect conc.:

1.1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

0.34 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Duration:

21 d

Dose descriptor:

LOEC

Effect conc.:

0.89 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Details on results:

The determined 21 day-LC50 and EC50 values were 1.0 mg/L and 1.1 mg/L based on the measured concentrations. The NOEC and LOEC for reproduction rate were evaluated to be 0.34 mg/L and 0.89 mg/L, respectively.

Results with reference substance (positive control):

The 48 h-EC50 (Immobility) value of the reference substance determined was 0.54 mg/L.

Reported statistics and error estimates:

The LC50 value was calculated with the Probit method. The EC50 value for reproduction rate was calculated with the Logistic method. For the determinations of NOEC and LOEC, the Dunnett method was used. A regression line for GC-MS analyses was derived by least square calculations. Numerical values were rounded according to the method specified in JIS.
The NOEC described above was defined as the maximum concentration at which reproduction rate was not significantly different from the control.
The LOEC was the minimum concentration at which significant difference was observed.

Cumulative number of dead daphnia and mortality

Measured Concentration [mg/L]	Day 21
	Cumulative Number of Dead	Mortality (%)
Control	0	0
Solvent Control	0	0
0.046	0	0
0.12	0	0
0.34	0	0
0.89	3	30
1.9	10	100

 

Mean cumulative numbers of juveniles produced per adult:

Measured Concentration [mg/L]	Mean cumulative Number of Juvenile (Day 21)
Control	101.9
Solvent Control	143.9
0.046	137.7
0.12	140.7
0.34	117.5
0.89	59.1
1.9	0.0

 

Validity criteria fulfilled:

yes

Remarks:

(- The immobilisation and other abnormalities in the controls did not exceed 10 % by the end of the test. - The dissolved oxygen concentration remained above 3 mg/L throughout the exposure period.)

Conclusions:

Daphnia magna was exposed under semi-static conditions for 21 days to five concentrations. The determined 21 day-LC50 and EC50 values were 1.0 mg/L and 1.1 mg/L based on the measured concentrations. The NOEC and LOEC for reproduction rate were evaluated to be 0.34 mg/L and 0.89 mg/L, respectively.

Executive summary:

The chronic toxicity of aquatic invertebrates, Daphnia magna, to 2,2'-methylenebis(6 -tert-butyl-4 -methylphenol) exposed to the test solution of five nominal concentrations (0.10, 0.22, 0.46, 1.0 and 2.2 mg/L), solvent control and control solution for a period of 21 days under semi-static conditins.

The study was conducted in accordance with OECD guideline No. 202 (April, 1984) and the revised version of OECD guideline no. 211 (April, 1997).

The determined 21 day-LC50 and EC50 values were 1.0 mg/L and 1.1 mg/L based on the measured concentrations. The NOEC and LOEC for reproduction rate were evaluated to be 0.34 mg/L and 0.89 mg/L, respectively. Experimental designs and results were well documented and prescribed conditions in the guideline were well satisfies.

Description of key information

The NOEC for reproduction was determined to be 0.34 mg/L. As the reported NOEC is above the water solubility no PNEC can be derived based on this result.

Additional information

The above study is assessed in the OECD SIDS Report (2001) and the PBT Report (2009) of the European Chemical Bureau (ECB). This test can be regarded as reliable without restrictions.

However the relevant test was performed with a solubilizer in order to overcome the low water solubility of 0.007 mg/L. As a consequence, no effects have been observed up to the limit of the water solubility. No PNEC can be derived based on this result.