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Short-term toxicity to aquatic invertebrates

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Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
24 February to 24 March 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
This study was performed according to OECD Guideline 202 and EU Method C.2 with GLP statement. All validity criteria were fulfilled.
Qualifier:
according to guideline
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes (incl. QA statement)
Remarks:
UK GLP Compliance Programme (inspected on July 05, 2016/ signed on October 28, 2016)
Specific details on test material used for the study:
- Stability in water: Stable (internal Firmenich data)
Analytical monitoring:
yes
Details on sampling:
- Concentrations: Samples were taken from the control and each test group from the bulk test preparation at 0 hours and from the pooled replicates at 48 hours for quantitative analysis.
- Sample storage conditions before analysis: Duplicate samples were taken at 0 and 48 hours and stored frozen for further analysis if necessary.
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: A nominal amount of test item (220 mg) was stirred with 2.2 liters of test water using a magnetic stirrer at a rate just fast enough to form a dimple at the water surface for 48 hours in a sealed vessel with minimal headspace. After the stirring period, the solution was settled for 24 hours before removing the aqueous phase avoiding any undissolved test item to produce a 100% v/v saturated solution of the test item. A series of dilutions was made from this saturated solution to give the test concentrations of 10, 5.6, 3.2, 1.8 and 1.0% v/v saturated solution. Each prepared concentration was inverted several times to ensure adequate mixing and homogeneity.
The concentration and stability of the test item in the test preparations was verified by chemical analysis at 0 and 48 hours
- Controls: The control group was maintained under identical conditions but not exposed to the test item.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Source: Test was carried out using first instar Daphnia magna derived from in-house laboratory cultures.
- Adult daphnids were maintained in 150 mL glass beakers containing Elendt M7 medium in a temperature controlled room maintaining the water temperature at 18-22 °C. The lighting cycle was controlled to give a 16 hours light and 8 hours darkness cycle with 20 minute dawn and dusk transition periods. Each culture was fed daily with a mixture of algal suspension (Desmodesmus subspicatus) and Tetramin® flake food suspension. Culture conditions ensured that reproduction was by parthenogenesis. Gravid adults were isolated the day before initiation of the test, such that the young daphnids produced overnight were less than 24 hours old. These young were removed from the cultures and used for testing.
- Age at study initiation: < 24 hours old
- Feeding during test: No feed was given during the exposure period.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Post exposure observation period:
None
Hardness:
No data
Test temperature:
21-22 °C
pH:
7.8-8.0 at the start (0 hours); 7.8-7.9 at termination (48 hours) of the test
Dissolved oxygen:
8.8-8.9 mg O2/L at the start (0 hours); 8.1-8.4 mg O2/L at termination (48 hours) of the test
Salinity:
None
Conductivity:
No data
Nominal and measured concentrations:
Nominal concentrations of 1.0, 1.8, 3.2, 5.6 and 10% v/v saturated solution
Details on test conditions:
TEST SYSTEM
- Test vessel: 100 mL conical flasks completely filled with test medium and sealed were used
- Type: Closed
- Test vessels were then covered to reduce evaporation
- Aeration: Test vessels were not aerated
- Renewal rate of test solution: Test preparations were not renewed during the exposure period
- 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 (Elendt M7) used for both the range-finding and definitive tests according to OECD 202

OTHER TEST CONDITIONS
- Photoperiod: 16 hours light and 8 hours darkness with 20 minute dawn and dusk transition periods
- Light intensity: 451 to 527 Lux

EFFECT PARAMETERS MEASURED
- Test Organism Observations: Any immobilization or adverse reactions to exposure were recorded at 24 and 48 hours after the start of exposure. The criterion of effect used was that daphnia were considered to be immobilized if they were unable to swim within 15 seconds after gentle agitation.
- Water Quality Criteria: Water temperature was recorded daily throughout the test. Dissolved oxygen concentrations and pH were recorded at the start and termination of the test. The pH and dissolved oxygen concentration were measured using a Hach Flexi handheld meter whilst the temperature was measured using a Hanna Instruments HI 93510 digital thermometer. The light intensity during the light period was measured using an ATP Instrumentation Lux meter. The appearance of the test media was recorded daily.

RANGE-FINDING STUDY
- Test concentrations: In the range-finding test Daphnia magna were exposed to a series of nominal test concentrations of 0.10, 1.0, 10 and 100% v/v saturated solution (10 daphnids per dose) for 48 hours.
- Results used to determine the conditions for the definitive study: No immobilization was observed at the test concentrations of 0.10 and 1.0% v/v saturated solution, however, immobilization was observed at 10 and 100% v/v saturated solution. Based on this information test concentrations of 1.0, 1.8, 3.2, 5.6 and 10% v/v saturated solution were selected for the definitive test.
Chemical analysis of the test preparations at 0 hours showed measured test concentrations to range from 0.041 to 44 mg/L. There was no significant change in the measured concentrations at 48 hours indicating that the test item was stable under test conditions.
Reference substance (positive control):
yes
Remarks:
potassium dichromate (Envigo Study Number XT01HS)
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
1.7 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95% Cl: 1.6-1.9 mg/L
Key result
Duration:
48 h
Dose descriptor:
other: Highest tested concentration without observed effect
Effect conc.:
1.2 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
Immobilization Data: No immobilization was observed at 24 hours; 0, 0, 0, 0, 80 and 100% immobilization was observed at 0, 0.36, 0.62, 1.2, 2.0 and 3.4 mg/L, respectively (48 hours).
Analysis of the immobilization data by the Trimmed Spearman-Karber procedure at 48 hours based on the 0-Hour measured test concentrations gave the following results: EC50 (24 h): > 3.4 mg/L; EC50 (48 h): 1.7 mg/L (95% Cl 1.6-1.9 mg/L).
The highest tested concentration without observed effect after 24 and 48 hours exposure were 3.4 and 1.2 mg/L respectively.
Sub-lethal effects: A sub-lethal effect of exposure was observed in the 3.4 mg/L test concentration at 24 hours and in the 2.0 mg/L test concentration at 48 hours. The response observed was reduced mobility.
Observations on test Item solubility: At the start and throughout the test all control and test solutions were observed to be clear colorless solutions.
Results with reference substance (positive control):
Potassium dichromate was used as the reference item at concentrations of 0.32, 0.56, 1.0, 1.8 and 3.2 mg/L (Envigo Study Number XT01HS). EC50 (24 h) and EC50 (48 h) were 1.3 and 1.2 mg/L, respectively.
The results from the positive control with potassium dichromate were within the normal range for this reference item (EC50-24 h within the range 0.6 to 2.1 mg/L).
Reported statistics and error estimates:
The EC50 value and associated confidence limits at 48 hours were calculated by the Trimmed Spearman-Karber procedure. The Lowest Observed Effect Concentration and the No Observed Effect Concentration at 24 hours were calculated using the Fisher’s Exact Binomial Test with Bonferroni correction and at 48 hours were calculated using the Step-down Cochran-Armitage Procedure. All results were calculated using the ToxRat Professional computer software package (TOXRAT).
An estimate of the 24 hour EC50 value was given by inspection of the immobilization data.

Table 6.1.3/1: Cumulative Immobilization Data and Observations in the Definitive Test

0 hour Measured Concentration (mg/L) 

Cumulative Immobilized Daphnia (Initial Population: 5 Per Replicate)

 

Observations

 

24 hours

Replicates

R1

R2

R3

R4

Total

%

R1

R2

R3

R4

Control

0

0

0

0

0

0

5 N

5 N

5 N

5 N

0.36

0

0

0

0

0

0

5 N

5 N

5 N

5 N

0.62

0

0

0

0

0

0

5 N

5 N

5 N

5 N

1.2

0

0

0

0

0

0

5 N

5 N

5 N

5 N

2.0

0

0

0

0

0

0

5 N

5 N

5 N

5 N

3.4

0

0

0

0

0

0

4 N, 1 R

5 N

5 N

5 N

48 hours

Control

0

0

0

0

0

0

5 N

5 N

5 N

5 N

0.36

0

0

0

0

0

0

5 N

5 N

5 N

5 N

0.62

0

0

0

0

0

0

5 N

5 N

5 N

5 N

1.2

0

0

0

0

0

0

5 N

5 N

5 N

5 N

2.0

5

4

4

3

16

80

A/I

1 R

1 R

2 R

3.4

5

5

5

5

20

100

A/I

A/I

A/I

A/I

 

R1 – R4 = Replicates 1 to 4

N = No sub-lethal effects observed

R = Reduced mobility

A/I = All daphnia immobilized

Verification of Test Concentrations

Chemical analysis of the test preparations at 0 hours showed measured test concentrations to range from 0.36 to 3.4 mg/L. There was no significant change in the measured concentrations at 48 and so the results are based on 0-Hour measured test concentrations only.

Analysis at 48 hours indicated the control media had been contaminated with test item. The measured concentration in the control at 48 hours was determined to be 0.090 mg/L. Given that the 0 hour control sample was shown not to be contaminated and that the test was conducted in completely filled and sealed test vessels it was considered that the contamination most likely occurred during sampling at the end of the daphnia exposure and therefore did not affect the results of the test.

Table 6.1.3/2: Measured concentrations

Time point

(hours)

Nominal concentrations

(% v/v saturated solution)

Measured concentrations

(mg/L)

0

Control

1.0

1.8

3.2

5.6

10

<LOQ

0.359

0.620

1.20

2.02

3.41

48

Control

1.0

1.8

3.2

5.6

10

0.0842 / 0.0967*

0.415

0.676

1.17

1.96

3.55

LOQ = Limit of Quantification

* = duplicate sample, stored frozen prior to analysis

Water Quality Criteria: Temperature was maintained at 21 °C to 22 °C throughout the test, while there were no treatment related differences for oxygen concentration or pH. Throughout the test the light intensity was observed to be in the range 451 to 527 Lux.

Validation Criteria: The test was considered to be valid given that none of the control daphnids showed immobilization or other signs of disease or stress and that the oxygen concentration at the end of the test was equal to or greater than 3 mg/L in the control and test vessels.

Validity criteria fulfilled:
yes
Conclusions:
Based on the 0 hour measured test concentrations, the 48-hour EC50 for test item was 1.7 mg/L (95% Cl 1.6-1.9 mg/L).
Executive summary:

The acute toxicity of the test item to Daphnia magna was assessed under static exposure conditions according to OECD Guideline 202 and EU Method C.2 with GLP statement.

Groups of twenty Daphnids, less than 24 hours old, were exposed to an aqueous solution of the test item at nominal concentrations of 1.0, 1.8, 3.2, 5.6 and 10% v/v saturated solution (initial measured concentrations of 0.36, 0.62, 1.2, 2.0 and 3.4 mg/L, respectively) for 48 hours at a temperature of 21-22 °C under static test conditions. Immobilization and any adverse reactions to exposure were recorded after 24 and 48 hours.

 

Chemical analysis of the test preparations at 0 hours showed measured test concentrations to range from 0.36 to 3.4 mg/L. There was no significant change in the measured concentrations at 48 hours and so the results are based on 0 hour measured test concentrations only.

 

No immobilization was observed at 24 hours; 0, 0, 0, 0, 80 and 100% immobilization was observed at 0, 0.36, 0.62, 1.2, 2.0 and 3.4 mg/L, respectively (48 hours). Analysis of the immobilization data by the Trimmed Spearman-Karber procedure at 48 hours based on the 0-Hour measured test concentrations gave the following results: EC50 (24 h): > 3.4 mg/L; EC50 (48 h): 1.7 mg/L (95% Cl 1.6-1.9 mg/L). The highest tested concentration without observed effect after 24 and 48 hours exposure were 3.4 and 1.2 mg/L respectively.

The results for the most recent test performed prior to this study indicated that, 24-hour EC50 for potassium dichromate (reference item) to Daphnia magna was 1.3 mg/L; this was within the range typically obtained in this laboratory (0.6 to 2.1 mg/L). Water quality parameters such as temperature, pH and dissolved oxygen were determined to be within the acceptable limits. All validity criteria were fulfilled.

 

Based on the 0 hour measured test concentrations, the 48-hour EC50 for test item was 1.7 mg/L (95% Cl 1.6-1.9 mg/L).

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
(Q)SAR
Adequacy of study:
supporting study
Study period:
Run on 2017/06
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
results derived from a valid (Q)SAR model and falling into its applicability domain, with limited documentation / justification
Justification for type of information:
1. SOFTWARE
ECOSAR

2. MODEL (incl. version number)
ECOSAR v1.11

3. SMILES OR OTHER IDENTIFIERS USED AS INPUT FOR THE MODEL
C1(CCC=C(C1)C(=O)CCC=C)C
log Kow = 4.1 (experimental result)
Water solubility = 56.2 mg/L (experimental result)

4. SCIENTIFIC VALIDITY OF THE (Q)SAR MODEL
- Defined Endpoint: short term toxicity to aquatic invertebrates (Daphnid).
- Unambigous algorithm: Linear regression QSAR; Log 48h-LC50 (mmol/L) = -0.858*log Kow + 1.3848. To convert the LC50 from mmol/L to mg/L, multiply by the molecular weight.
- Defined domain of applicability: Applicable to chemicals with log Kow less than 5.0 and molecular weight less than 1000 g/mol. The model can be suitable for chemicals class of Neutral organics.
- Appropriate measures of goodness-of-fit and robustness and predictivity: N = 98+31 and the coefficient of determination R² = 0.7704.
- Mechanistic interpretation: related to the partitioning of the substance from water into the organisms lipid phase.

5. APPLICABILITY DOMAIN
Applicable to chemicals with log Kow less than 5.0 and molecular weight less than 1000 g/mol. The model can be suitable for chemicals class of Neutral organics.

6. ADEQUACY OF THE RESULT
The substance falls within the applicability domain described above and therefore the predicted value can be considered reliable.
For more information, see Neutral organics SAR document in "Attached background material".
Qualifier:
according to guideline
Guideline:
other: REACH guidance on QSARs R.6, May 2008
Deviations:
no
Principles of method if other than guideline:
See attached QPRF.
GLP compliance:
no
Remarks:
(not relevant)
Specific details on test material used for the study:
No additional information
Details on sampling:
Not applicable
Details on test solutions:
Not applicable
Test organisms (species):
Daphnia sp.
Details on test organisms:
None
Test type:
not specified
Water media type:
not specified
Total exposure duration:
48 h
Remarks on exposure duration:
none
Post exposure observation period:
Not applicable
Hardness:
Not applicable
Test temperature:
Not applicable
pH:
Not applicable
Dissolved oxygen:
Not applicable
Salinity:
Not applicable
Nominal and measured concentrations:
Not applicable
Details on test conditions:
Not applicable
Key result
Duration:
48 h
Dose descriptor:
LC50
Effect conc.:
1.312 mg/L
Remarks on result:
other: ECOSAR Class: Neutral Organics (baseline toxicity)
Duration:
48 h
Dose descriptor:
LC50
Effect conc.:
1.569 mg/L
Remarks on result:
other: ECOSAR Class: Vinyl/Allyl Ketones
Details on results:
Two ECOSAR class are available for the prediction. The worst case model was selected as key result.

Validity of model:
1. Defined Endpoint: short term toxicity to aquatic invertebrates (Daphnid).
2. Unambigous algorithm: Linear regression QSAR; Log 48h-LC50 (mmol/L) = -0.858*log Kow + 1.3848.
To convert the LC50 from mmol/L to mg/L, multiply by the molecular weight.
3. Applicability domain: applicable to chemicals with log Kow less than 5.0 and molecular weight less than 1000 g/mol. The model can be suitable for chemicals class of neutral organic.
4. Statistical characteristics: N = 98+31 and the coefficient of determination R² = 0.7704.
5. Mechanistic interpretation: related to the partitioning of the substance from water into the organisms lipid phase.

Adequacy of prediction: the substance falls within the applicability domain described above and therefore the predicted value can be considered reliable. For more information, see Neutral Organics SAR document in "Attached background material".
Results with reference substance (positive control):
Not applicable
Reported statistics and error estimates:
N = 98+31 and the coefficient of determination R² = 0.7704.

None

Validity criteria fulfilled:
yes
Conclusions:
The substance was predicted to have a LC50 value at 1.312 mg/L to aquatic invertebrates Daphnid after 48h of exposure. The substance falls within the applicability domain and therefore the predicted value can be considered reliable.
Executive summary:

The short term toxicity to daphnid was estimated using the ECOSAR v1.11 QSAR model available from the U.S. EPA.

The estimated 48h-LC50 value was 1.312 mg/L (Neutral Organics class), calculated from a measured log Kow value at 4.1, a measured water solubility value at 56.2 mg/L and a linear regression:

Log 48h-LC50 (mmol/L) = -0.858*log Kow + 1.3848.

The substance falls within the applicability domain and therefore the predicted value can be considered reliable.

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
(Q)SAR
Adequacy of study:
supporting study
Study period:
Run on 2017/06
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
results derived from a valid (Q)SAR model and falling into its applicability domain, with limited documentation / justification
Justification for type of information:
1. SOFTWARE
ECOSAR

2. MODEL (incl. version number)
ECOSAR v1.11

3. SMILES OR OTHER IDENTIFIERS USED AS INPUT FOR THE MODEL
CC1(C)CCCC(=O)C1
log Kow = 2.0 (experimental result)

4. SCIENTIFIC VALIDITY OF THE (Q)SAR MODEL
- Defined Endpoint: short term toxicity to aquatic invertebrates (Daphnid).
- Unambigous algorithm: Linear regression QSAR; Log 48h-LC50 (mmol/L) = -0.858*log Kow + 1.3848. To convert the LC50 from mmol/L to mg/L, multiply by the molecular weight.
- Defined domain of applicability: Applicable to chemicals with log Kow less than 5.0 and molecular weight less than 1000 g/mol. The model can be suitable for chemicals class of Neutral organics.
- Appropriate measures of goodness-of-fit and robustness and predictivity: N = 98+31 and the coefficient of determination R² = 0.7704.
- Mechanistic interpretation: related to the partitioning of the substance from water into the organisms lipid phase.

5. APPLICABILITY DOMAIN
Applicable to chemicals with log Kow less than 5.0 and molecular weight less than 1000 g/mol. The model can be suitable for chemicals class of Neutral organics.

6. ADEQUACY OF THE RESULT
The substance falls within the applicability domain described above and therefore the predicted value can be considered reliable.
For more information, see Neutral organics SAR document in "Attached background material".
Qualifier:
according to guideline
Guideline:
other: REACH guidance on QSARs R.6, May 2008
Deviations:
no
Principles of method if other than guideline:
See attached QPRF.
GLP compliance:
no
Remarks:
(not relevant)
Specific details on test material used for the study:
No additional information
Details on sampling:
Not applicable
Details on test solutions:
Not applicable
Test organisms (species):
Daphnia sp.
Details on test organisms:
None
Test type:
not specified
Water media type:
not specified
Total exposure duration:
48 h
Remarks on exposure duration:
none
Post exposure observation period:
Not applicable
Hardness:
Not applicable
Test temperature:
Not applicable
pH:
Not applicable
Dissolved oxygen:
Not applicable
Salinity:
Not applicable
Nominal and measured concentrations:
Not applicable
Details on test conditions:
Not applicable
Key result
Duration:
48 h
Dose descriptor:
LC50
Effect conc.:
58.865 mg/L
Remarks on result:
other: ECOSAR Class: Neutral Organics (baseline toxicity)
Details on results:
Validity of model:
1. Defined Endpoint: short term toxicity to aquatic invertebrates (Daphnid).
2. Unambigous algorithm: Linear regression QSAR; Log 48h-LC50 (mmol/L) = -0.858*log Kow + 1.3848.
To convert the LC50 from mmol/L to mg/L, multiply by the molecular weight.
3. Applicability domain: applicable to chemicals with log Kow less than 5.0 and molecular weight less than 1000 g/mol. The model can be suitable for chemicals class of neutral organic.
4. Statistical characteristics: N = 98+31 and the coefficient of determination R² = 0.7704.
5. Mechanistic interpretation: related to the partitioning of the substance from water into the organisms lipid phase.

Adequacy of prediction: the substance falls within the applicability domain described above and therefore the predicted value can be considered reliable. For more information, see Neutral Organics SAR document in "Attached background material".
Results with reference substance (positive control):
Not applicable
Reported statistics and error estimates:
N = 98+31 and the coefficient of determination R² = 0.7704.

None

Validity criteria fulfilled:
yes
Conclusions:
The substance was predicted to have a LC50 value at 58.865 mg/L to aquatic invertebrates Daphnid after 48h of exposure. The substance falls within the applicability domain and therefore the predicted value can be considered reliable.
Executive summary:

The short term toxicity to daphnid of 3,3-dimethyl cyclohexanone (DMCH), the relevant degradation product of the registered substance, was estimated using the ECOSAR v1.11 QSAR model available from the U.S. EPA.

The estimated 48h-LC50 value was 58.865 mg/L (Neutral Organics class), calculated from a measured log Kow value at 2.0 and a linear regression:

Log 48h-LC50 (mmol/L) = -0.858*log Kow + 1.3848.

The substance falls within the applicability domain and therefore the predicted value can be considered reliable.

Description of key information

OECD Guideline 202, EU Method C.2, GLP, key study, validity 1:

48h-EC50 (Daphnia magna) = 1.7 mg/L (95% CL: 1.6 - 1.9 mg/L) based on the initial measured concentrations.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Effect concentration:
1.7 mg/L

Additional information

To assess the short-term toxicity of the registered substance to aquatic invertebrates, one valid experimental study and a QSAR prediction are available.

In the experimental study (Envigo, 2017), assessed as the key study, the acute toxicity of the registered substance to Daphnia magna was assessed under static exposure conditions according to OECD Guideline 202 and EU Method C.2 with GLP statement. Groups of twenty Daphnids (less than 24 hours old) were exposed to an aqueous solution of the test item at nominal concentrations of 1.0, 1.8, 3.2, 5.6 and 10% v/v saturated solution (initial measured concentrations of 0.36, 0.62, 1.2, 2.0 and 3.4 mg/L, respectively) for 48 hours at a temperature of 21 -22 °C under static test conditions. Immobilization and any adverse reactions to exposure were recorded after 24 and 48 hours. Chemical analysis of the test preparations at 0 hours showed measured test concentrations to range from 0.36 to 3.4 mg/L. There was no significant change in the measured concentrations at 48 hours and so the results are based on 0 hour measured test concentrations only. According to the results of this study, the 48h-EC50 value was determined at 1.7 mg/L (95% CL: 1.6-1.9 mg/L), based on the initial measured concentrations.

In addition, to support the QSAR approach used for the short-term toxicity to fish endpoint, a QSAR prediction (ECOSAR, 2017), assessed as a supporting data, was provided. This QSAR prediction was performed on the registered substance to assess the short-term toxicity of the substance to daphnids, using the ECOSAR v1.11 QSAR model available from the U.S. EPA.

The estimated 48h-LC50 value was 1.31 mg/L (Neutral Organics class), calculated from a measured log Kow value at 4.1, a measured water solubility value at 56.2 mg/L and a linear regression. The substance falls within the applicability domain and therefore the predicted value can be considered reliable.

This QSAR result supports the key study performed on the same substance with a similar endpoint value, slightly more conservative, validating the model for use with this substance.

In conclusion, according to the key experimental study (Envigo, 2017), the 48h-EC50 value on Daphnia magna is 1.7 mg/L (95% CL: 1.6 - 1.9 mg/L).

Moreover, as the registered substance is totally primarily biodegraded in the environment to form two degradation products: succinic acid (degraded also and mineralized completely) and 3,3 -dimethyl cyclohexanone (DMCH), the short-term toxicity of the relevant degradation product of the registered substance, DMCH, to daphnids was estimated using the same QSAR model presented above, ECOSAR v1.11 available from the U.S. EPA. The estimated 48h-LC50 value was 58.87 mg/L (Neutral Organics class), calculated from a measured log Kow value at 2.0 and a linear regression. The substance falls within the applicability domain and therefore the predicted value can be considered reliable. According to this result, the relevant degradation product of the registered substance, DMCH, is presumed not Toxic (T) in the PBT assessment context.