Registration Dossier

Ecotoxicological information

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:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018
Report Date:
2018

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Version / remarks:
2008
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Version / remarks:
adopted April 13, 2004
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
- Concentrations: 0, 100 mg/L
- Sample storage conditions before analysis: Routinely, the samples were analysed immediately. Only in exceptional cases, they were stored overnight deep frozen and protected from light.

Test solutions

Vehicle:
no
Remarks:
medium was used
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- To produce the only test item concentration, 100.9 mg of the test item were added to 1 litre of dilution water, treated for 1 h in an ultrasonic bath and stirred for 24 h on a magnetic stirrer. Undissolved particles of the test item were removed by filtration using a folded filter with a pore size of 7 - 12 µm. The pH was measured to be 5.9 and was adjusted to pH 6.2.
50 mL of the solution were taken and diluted with 0.5 mL of dilution water containing 5 daphnids resulting in a final concentration of 100 mg/L. For each test item concentration and the control 4 replicates were prepared.
- Controls: medium (composition see attachment 3) without any test item
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): Undissolved particles of the test item were removed by filtration using a folded filter with a pore size of 7 - 12 µm.

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Source: Strain of Bundesgesundheitsamt Berlin
- Maintenance and Acclimatisation : A population of parthenogenetic females of synchronized age structure has been maintained for more than 20 years in the test facility under constant temperature conditions (20 +/- 2 °C) at a 16 : 8 hour light-dark photoperiod (light intensity:
< 20 µE x m-2 x s-1). The culture water (so-called 'M4 medium') was partly renewed once a week. The Daphnia were exclusively fed unicellular green algae (Desmodesmus subspicatus) 'ad libitum'. Mortalities of parent Daphnia during the culture period were recorded daily. The neonates were separated from their parent Daphnia by filtration prior to the acute test. No feeding during the exposure period.

Study design

Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
48 h

Test conditions

Hardness:
The hardness of the dilution water used was 14.5 °dH (= 259 mg/L CaCO3).
Test temperature:
19.3 to 19.6 °C
pH:
The pH was measured to be 5.9 and was adjusted to pH 6.2.
Dissolved oxygen:
8.7 - 8.8 mg O2/L
Nominal and measured concentrations:
Nominal: 100 mg/L
Measured: 103.838 mg/L after 0 h, 103.822 mg/L after 48 h
Details on test conditions:
TEST SYSTEM
- Test vessel, Type: 100 mL glass beakers covered with watch glasses holding 5 neonates in 50 mL of test medium
- Aeration: none
- 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: Reconstituted water (so-called 'M4 medium' according to OECD 202 and EC Method C.2, attachment 3) was used for the maintenance of the test animals and the preparation of stock and test solutions of the test item.

OTHER TEST CONDITIONS
- Adjustment of pH: yes
- Photoperiod: 16 h light : 8 h dark

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : The criterion of adverse effects used in this study was the item-induced alteration of the normal mobility behaviour and the loss of locomotory actions of the neonates, observed at 24 and 48 hours.

VEHICLE CONTROL PERFORMED: yes, medium only

RANGE-FINDING STUDY
- Test concentrations: A range finding test preceded the main test. It provided information about the range of concentrations which were used in the main test. The following nominal concentrations of the test item were tested in the range finding test: 1, 10 and 100 mg/L.
Reference substance (positive control):
no

Results and discussion

Effect concentrationsopen allclose all
Duration:
24 h
Dose descriptor:
EC0
Effect conc.:
>= 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Duration:
24 h
Dose descriptor:
EC50
Effect conc.:
> 100 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:
EC0
Effect conc.:
>= 100 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mobility
Key result
Duration:
48 h
Dose descriptor:
EC50
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

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
The study was conducted under GLP according to EU method C.2, which is equivalent to OECD guideline 202, on the registered substance itself. The method is to be considered scientifically reasonable with no deficiencies in documentation or deviations from the guidelines, the validity criteria were met. Hence, the results can be considered as reliable to assess the toxicity of the test substance towards daphnids.
The toxicity against Daphnia magna was tested in a static limit test at a nominal test concentration of 100 mg/L. The concentrations showed no toxicity. None of the animals were immobilised neither in the treatment nor in the control.
Based on the obtained results, the substance does not need to be classified as hazardous to the aquatic environment according to the Regulation (EC) No. 1272/2008.
Executive summary:

A GLP study was performed to assess the acute toxicity of Hydroxytoluic acid to Daphnia magna STRAUS under static conditions.

The study was conducted in accordance with the Council Regulation (EC) No 440/2008, Method C.2 ‘Acute toxicity for Daphnia’ (2008) which is equivalent to OECD Guideline for Testing of Chemicals No. 202 'Daphnia sp., Acute Immobilisation Test' (adopted April 13, 2004).

The Daphnia were exposed to a limit test concentration of nominally 100 mg/L of Hydroxytoluic acid dissolved in dilution water. Auxiliaries used to prepare the test media were an ultrasonic bath, a magnetic stirrer and a folded filter.

Observations were made on the swimming ability and the immobilisation rate, respectively, after 24 and 48 hours of exposure. The following values were determined:

Time

[h] 

 EC50

[mg/L]

 24  > 100
 48  > 100

No toxic effects against Daphnia were observed at a limit test concentration of 100 mg/L under exposure conditions.

The results are expressed in terms of nominal concentrations. Effective concentrations correspond to 103.8% of nominal values at 0 hours, and to 103.8% of nominal values at 48 hours. The hardness of the dilution water used was 14.5°dH (= 259 mg/L CaCO3).