6,8-dimethylnon-7-enal

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

(Q)SAR

Adequacy of study:

key study

Study period:

October 18th, 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

results derived from a valid (Q)SAR model and falling into its applicability domain, with adequate and reliable documentation / justification

Justification for type of information:

1. SOFTWARE
iSafeRat® High-Accuracy-Quantitative Structure-Activity Relationship (HA-QSAR) based on a holistic approach for predicting physicochemical and ecotoxicological endpoints: Short-term toxicity to Daphnia (immobilisation)

2. MODEL (incl. version number)
iSafeRat® holistic HA-QSAR v1.8

3. SMILES OR OTHER IDENTIFIERS USED AS INPUT FOR THE MODEL
SMILES: O=CCCCCC(C)C=C(C)C
The toxicity of the test item was predicted using the iSafeRat® Ecotox module providing the Subcooled Liquid Water Solubility (SLWS) as the input. The SLWS has been predicted using the iSafeRat® Water Solubility module providing the experimental log KOW value as the input. Water Solubility of the test item = 40.2 mg/L (or -3.622 in log (mol/L).

4. SCIENTIFIC VALIDITY OF THE (Q)SAR MODEL
See attached QMRF

5. APPLICABILITY DOMAIN
See attached QPRF

6. ADEQUACY OF THE RESULT
See attached QPRF

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

not applicable

Remarks:

QSAR model

Principles of method if other than guideline:

The purpose of this QSAR model is to accurately predict the acute toxicity to daphnids as would be expected in a laboratory experiment following OECD Guideline 202 and EC method C.2 for specific, named mechanisms of action. The model provides an in silico prediction for the 48-hour EC50 value that can effectively be used in place of an experimentally derived 48-hour EC50 value. The regression based method used to achieve this has been fully validated following the OECD recommendations (OECD, 2004).

GLP compliance:

no

Remarks:

QSAR model

Specific details on test material used for the study:

- log KOW = 3.87 (Phytosafe, 2008, EU Method A.8, Flask Method)
- Water Solubility 40.2 mg/L (KREATiS, 2019)
- Mechanism of action: MechoA 3.1: hard electrophiles (Bauer et al., 2018)

Analytical monitoring:

no

Details on sampling:

not applicable

Vehicle:

no

Details on test solutions:

not applicable

Test organisms (species):

other: Daphnia magna, Daphnia pulex

Details on test organisms:

No difference in terms of toxic mechanism of action between invertebrate (or indeed other) aquatic species is expected. Any observed differences may be attributed to lifestyle related parameters (e.g. shell closing in molluscs) and relative duration of study versus bodysize rather than to a specific toxic mechanism causing species differences.

Test type:

other: QSAR model

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Remarks on exposure duration:

Results from a test duration of 48 hours only were used for daphnid species.

Post exposure observation period:

not applicable

Hardness:

The QSAR is based on data from studies performed at acceptable hardness to ensure control survival.

Test temperature:

The temperatures varied from approximately 20 to 23 °C depending on the species used to construct the algorithm. This small difference is not expected to contribute to the variability of the EC50 values found in experimental data.

pH:

Test results were taken from studies with measured pHs between 6 - 9.

Dissolved oxygen:

The model is based on data from studies performed at acceptable oxygen concentrations (generally >60%).

Salinity:

Not applicable

Conductivity:

Not applicable

Nominal and measured concentrations:

Studies were used only where sufficient evidence was presented to determine that the substance was stable under test conditions (i.e. maintained within ± 20 % of the nominal) or, if not, the result was based on measured concentrations as geometric mean.

Details on test conditions:

Preferentially results from semi-static studies were used. However, substances tested using a static design were accepted (preferably accompanied by analytical measurements over the study period). For suspected volatile substances only tests performed in closed vessels were accepted unless accompanying analytical monitoring proved such a design was not necessary.

Reference substance (positive control):

no

Remarks:

QSAR model

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

1.2 mg/L

Nominal / measured:

meas. (not specified)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95%CL: 0.84 - 1.6 mg/L

Details on results:

The test item falls within the applicability domain of the model and was therefore reliably predicted for its acute toxicity (48h-EC50) to daphnids. Therefore, this endpoint value can be considered valid for use in risk assessment and classification and labelling.
The 48h-EC50 of the test item to daphnids was predicted as 1.2 mg/L.

Results with reference substance (positive control):

not applicable

Reported statistics and error estimates:

95% confidence interval (α = 0.05): 0.84 – 1.6 mg/L.
QSAR statistical parameters are given in the QMRF and the QPRF

Applicability Domain of the model

Descriptor domain:

The Subcooled Liquid Water Solubility value (40.2 mg/L or -3.622 in log (mol/L)) given as the input to the Ecotox module of the iSafeRat® Holistic HA-QSAR falls within the descriptor domain of the model between a log water solubility (in log (mol/L)) of -5.15 to -0.080.

Structural fragment domain:

All chemical groups within the molecular structure are taken into account by the model.

Mechanistic domain:

Currently, the ecotoxicity module of the iSafeRat® Holistic HA-QSAR can reliably predict the aquatic toxicity for

chemicals with the following mechanisms of action of toxicity (MechoA):

• non-polar narcosis (MechoA 1.1)

• polar narcosis of alkyl-/alkoxy-phenols (MechoA 1.2)

• polar narcosis of aliphatic amines (MechoA 1.2)

• cationic narcosis of quaternary ammoniums (MechoA 1.3)

• mono-/poly-esters whose hydrolysis products are narcotics (MechoA 2.1)

• hard electrophile reactivity (MechoA 3.1)

• RedOx cycling of primary thiols (MechoA 4.4)

• Proton release of carboxylic acids (MechoA 5.2)

The MechoA of molecules is predicted directly from the structure. The test item as an aldehyde is expected to exert a MechoA 3.1 and can be taken into account by the model (Bauer et al., 2018).

See QMRF in "attached background material".

Validity criteria fulfilled:

yes

Conclusions:

The test item falls within the applicability domain of the model and was therefore reliably predicted for its acute toxicity (48h-EC50) to daphnids. Therefore, this endpoint value can be considered valid for use in risk assessment and classification and labelling.
The 48h-EC50 of the test item to daphnids was predicted as 1.2 mg/L.
95% confidence interval (α = 0.05): 0.84 - 1.6 mg/L.

Executive summary:

A Quantitative Structure-Activity Relationship (QSAR) model was used to calculate the acute toxicity of the test item to daphnid. This QSAR model has been validated to be compliant with the OECD recommendations for QSAR modeling (OECD, 2004a) and predicts the endpoint value which would be expected when testing the substance under experimental conditions in a laboratory following the Guideline for Testing of Chemicals No. 202, "Daphnia sp., Acute Immobilisation Test" (OECD, 2004b), referenced as Method C.2 of Commission Regulation No. 440/2008 (European Commission, 2008). The criterion predicted was the EC50 (Median Effective Concentration), a statistically derived concentration which is expected to cause immobility in 50% of test animals within a period of 48 hours.

The immobility of the daphnids was determined using a validated QSAR model for the Mechanism of Action (MechoA) in question (MechoA 3.1, i.e. hard electrophiles as aldehyde) (Bauer et al., 2018).

The QSAR model is based on validated data for a small but fully validated training set of chemicals derived from 48-hour test on daphnids, for which the concentrations of the test item had been determined by chemical analyses over the test period.The correlation coefficient R2 and Root Mean Standard Error (RMSE) values for this model were 0.9913 and 0.0700, respectively.

The 48h-EC50 of the test item to daphnids was predicted as 1.2 mg/L.

95% confidence interval (α = 0.05): 0.84 - 1.6 mg/L.

Reference 2

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

disregarded due to major methodological deficiencies

Study period:

2010

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

significant methodological deficiencies

Remarks:

This study was performed according to OECD Guideline 202 with GLP certificate. All validity criteria were fulfilled. However, this study is not reliable due to high concentration of solvent used. Indeed, a large quantity of acetone was used as solvent in this study. Because of the potential for interaction with the test chemical resulting in an altered response in the test, solvent use should be restricted to situations where no other acceptable method of test solution preparation is available. The use of solvent is not the best method in the case of moderately soluble substances such as this one. Considering the acceptably high water solubility of the substance (40,2 mg/L) and the concentrations used in this study, this method could have been avoided. Furthermore, solvents are generally not appropriate for multiconstituent substances, like the test substance (which is a mixture of isomers), where the use of the solvent can give preferential dissolution of one or more components and thereby affect the toxicity. Then, the concentration/quantity of solvent used in the treatment solutions was 5 mL/L , corresponding to 3.95 g/L (with a density of 0.79), which is 50 times higher than the recommended maximum level of solvent (below 0.1 mL/L; OECD No. 23) and represent almost half of the EC50 of acetone (which was reported in the ECHA disseminated dossier at 8.8 g/L). Acetone is suspected to have affected the definitive results. Some solvent/substance interactions may have occurred in this study with this high concentration of acetone.

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

April 2004

Deviations:

yes

Remarks:

M4 mineral medium was used instead of ISO 6341 reconstituted water ; the range-finding test involved 2 replicate units for reach test item treatment and for the controls instead of one. These deviations did not adversely affect the validity of the study.

GLP compliance:

yes (incl. QA statement)

Remarks:

GLP Compliance statement signe on October 6th, 2010

Specific details on test material used for the study:

- Storage condition of test material: Stored at room temperature protected from direct sunlight

Analytical monitoring:

yes

Remarks:

The test item was quantifies for each test item concentration, at test initation, and at the end of the test, and at the medium renewal, in the old and new medium (definitive test).

Details on sampling:

- Concentrations: The test item was quantifies for each test item concentration, at test initation, and at the end of the test, and at the medium renewal, in the old and new medium (definitive test).
- Sampling method: At each the first and the second period, duplicate set of test tubes were prepared. The first one was used for the quantification of the test item at test initiation, or in the new medium (after 24h of testing). The second one was kept under the same environmental conditions as the test tubes and was used for the quantification of the test item at the end of the test, or in the old medium (after 24h of testing).

Vehicle:

yes

Remarks:

Acetone is used as solvent in the test.

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The treatment solutions were preapred in acetone. The test item concentrations were set at 120% of the nominal azlues using 50µL for each treatment solution, resulting in 1.0, 1.7, 2.9, 4.9 and 8.3 mg/L test item treatments (nominal values), respectively. The test solutions were renewed after 24h of testing. The daphnids were transferred to new test tubes filled with the new test solutions, and the controls (water and solvent) were transferred similarly.
- Controls: water control : the control tubes were made with 10 ML of reconstituted water ; solvent control : 10 mL of reconstituted water and 50 µL of acetone
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): acetone
- Concentration of vehicle in test medium: 5 mL/L

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnid
- Strain/clone: Daphnia magna
- Source: the strain was received from a commercial breeder and regularly sub-cultured at the phytosafe site.
- Age of parental stock (mean and range, SD): Adult females were transferred in a new volume of water on the day before the start of the test. Young daphnids produced within 24 hours were used for the tests.
- Feeding during test: Daphnids were no fed during the exposure period.
- Food type: Living cells of the green algae Desmodesmus subspicatus were provided as food.

Test type:

semi-static

Water media type:

other: The M4 mineral medium was used instead of ISO 6341 reconstituted water. The mineral medium was freshly reconstituted from analytical water and aliquot samples of mineral stock solutions.

Limit test:

no

Total exposure duration:

48 h

Test temperature:

20,0 +/- 1,0 °C

pH:

from 8.04 to 8.76

Dissolved oxygen:

from 6.8 to 8.5 mg Dissolved O2/L

Nominal and measured concentrations:

- Nominal concentrations : 1,0 ; 1,7 ; 2,9 ; 4,9 and 8,3 mg/L.
- Measured concentrations: see table 2 in "Any other information on results incl. tables"

Details on test conditions:

TEST SYSTEM
- Test vessel: 15 mL capacity glass tubes filles with 10 mL or reconstituted water
- Type (delete if not applicable): closed
- Rewal : The definitive test was performed using semi-static methodology: the test item treatments were set at 120% of the nominal values and the treatment solutions were renewed after 24h of testing so that the susbtance concentrations were maintained within +/- 20% of the nominal value throughout the test.
- No. of organisms per vessel: 5 daphnids per unit
- No. of vessels per concentration (replicates): 4 replicates per concentration
- No. of vessels per control (replicates): 4 replicate units
- No. of vessels per vehicle control (replicates): 4 replicate units

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The M4 mineral medium was used instead of ISO 6341 reconstituted water. The mineral medium was freshly reconstituted from analytical water and aliquot samples of mineral stock solutions.

OTHER TEST CONDITIONS
- Photoperiod: the tubes were maintained at dark in a climatic chamber.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Each test tube was checked for immobilized daphnids, 24 and 48 hours after the start of the test. Any abnormal behavior or appearance, if any, was reported.

VEHICLE CONTROL PERFORMED: yes

RANGE-FINDING STUDY
- Test concentrations: nominal : 0,01 ; 0,1 ; 1,0 ; 10,0 and 100,1 mg/L
- Results used to determine the conditions for the definitive study: No significant immobilization was observed for the three lowest test item treatments after 48h : 0% immoblization observed between 0,01 and 1,0 mg/L ; Daphnids were immobilized at both 10,0 and 100,1 mg/L : 100% immobilization at 10,0 mg/L and 100,1 mg/L was observed after 48h of exposure.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

2.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% CI = 1,4-4,3 mg/L

Results with reference substance (positive control):

The 24h-EC50 for Potassium dichromate was between 0,6 and 1,7 mg/L.

Table 1 : Percentage immbilization for the definitive test

	% immoibilization after 24h				% immobilization after 48h
Water control	0	0	0	0	0	0	0	0
Solvent control	0	0	0	0	0	0	0	0
Test substance
1,0 mg/L	20	0	0	0	20	0	0	0
1,7 mg/L	0	0	20	0	40	20	20	20
2,9 mg/L	20	20	0	0	40	80	40	60
4,9 mg/L	40	40	20	60	100	100	60	100
8,3 mg/L	40	80	100	80	60	100	100	100

Table 2: Analytical measurements for the definitive test

Measured concentrations	Nominal concentrations – mg/L
	1.01	1.71	2.90	4.92	8.34
T0, Test initiation (% nominal value)	1.22 (120.8%)	1.97 (114.9%)	3.018 (103.7%)	5.21 (105.8%)	8.97 (107.6%)
T0+24h, Old medium (% nominal value)	0.83 (82.4%)	1.38 (80.8%)	2.83 (97.5%)	4.85 (98.5%)	8.00 (95.9%)
T0+24h, New medium (% nominal value)	1.19 (118.2%)	1.98 (115.9%)	3.35 (115.4%)	5.42 (110.1%)	9.27 (111.1%)
T48, End of test (% nominal value)	1.15 (113.6%)	1.92 (112.3%)	3.19 (109.9%)	5.38 (109.3%)	9.10 (109.1%)

Validity criteria fulfilled:

yes

Remarks:

Immobilization was less than 10% in the control tubes, the dissolved oxygen concentration exceeded 75% in the control and test vessels, the test item concentrations were satisfactorily maintained between 80-120 % the nominal values over the test period.

Conclusions:

The 48h-EC50 was determined at 2,5 mg/L with a 95%-confidence interval of 1,4 -4,3 mg/L, based on analytically confirmed nominal concentrations.
Even if all validity criteria were fulfilled, this study is not reliable due to high concentration of solvent used.

Executive summary:

The influence of the test substance on mobility of Daphnia magna was investigated according to OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test) (April 2004), with GLP compliance statement. Daphnia were exposed to the test item under semi-static conditions, for 48 h.

Based on the results of a range-finding test, a definitive test was performed, exposing Daphnia magna to 5 different nominal concentrations : 1,0 ; 1,7 ; 2,9 ; 4,9 and 8,3 mg/L. The test item was quantified for each concentration at test initiation and at the end of test and at the medium renewal, in the old and new medium (definitive test). Each test tube was checked for immobilized daphnids 24 and 48 hours after the start of the test.

The 48h-EC50 was determined at 2,5 mg/L with a 95%-confidence interval of 1,4 -4,3 mg/L, based on analytically confirmed nominal concentrations.

Even if all validity criteria were fulfilled, this study is not reliable for the following reasons:

- Acetone was used as solvent in this study. Because of the potential for interaction with the test chemical resulting in an altered response in the test, solvent use should be restricted to situations where no other acceptable method of test solution preparation is available.  The use of solvent is not the best method at the time being.

- Considering the acceptably high water solubility of the substance (40,2 mg/L) and the concentrations used in this study, this method could have been avoided.

- Solvents are generally not appropriate for multiconstituent substances, like the test substance (which is a mixture of isomers), where the use of the solvent can give preferential dissolution of one or more components and thereby affect the toxicity.

- The concentration/quantity of solvent used in the treatment solutions was 5 mL/L , corresponding to 3.95 g/L (with a density of 0.79), which is 50 times higher than the recommended maximum level of solvent (below 0.1 mL/L; OECD No. 23) and represent almost half of the EC50 of acetone (which was reported in the ECHA disseminated dossier at 8.8 g/L). Acetone is suspected to have affected the definitive results. Some solvent/substance interactions may have occurred in this study with this high concentration of acetone.

Description of key information

iSafeRat® High-Accuracy-Quantitative Structure-Activity Relationship, KREATIS, 2019 :

48h-EC50 = 1.2 mg/L (95% confidence interval: 0.84 – 1.6 mg/L)

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

1.2 mg/L

Additional information

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

The experimental study (Phytosafe, 2010) was considered as not reliable and was disregarded due to major methodological deficiencies. Indeed, a large quantity of Acetone was used as solvent in this study. Because of the potential for interaction with the test chemical resulting in an altered response in the test, solvent use should be restricted to situations where no other acceptable method of test solution preparation is available. The use of solvent is not the best method in the case of moderately soluble substances such as this one. Considering the acceptably high water solubility of the substance (40.2 mg/L) and the test concentrations used in this study, this method could have been avoided. Furthermore, solvents are generally not appropriate for multiconstituent substances, such as this test substance (which is a mixture of isomers), where the use of the solvent can give preferential dissolution of one or more components and thereby affect the toxicity. The concentration/quantity of solvent used in the treatment solutions was 5 mL/L , corresponding to 3.95 g/L (with a density of 0.79), which is 50 times higher than the recommended maximum level of solvent (below 0.1 mL/L; OECD No. 23) and represent almost half of the EC50 of acetone (which was reported in the ECHA disseminated dossier at 8.8 g/L). Therefore, solvent/substance interactions may have occurred in this study with this high concentration of acetone and the result cannot be considered fit for use.

The QSAR prediction (KREATiS, 2019) was considered as reliable. The QSAR model has been validated to be compliant with the OECD recommendations for QSAR modeling (OECD, 2004) and predicts the endpoint value which would be expected when testing the substance under experimental conditions in a laboratory following the OECD Guideline 202. The immobility of the daphnids was determined using a validated QSAR model for the Mechanism of Action (MechoA) in question (MechoA 3.1, i.e. hard electrophiles as aldehyde) (Baueret al.,2018). The QSAR model is based on validated data for a small but fully validated training set of chemicals derived from 48-hour test on daphnids, for which the concentrations of the test item had been determined by chemical analyses over the test period. The correlation coefficient R2 and Root Mean Standard Error (RMSE) values for this model were 0.9913 and 0.0700, respectively.

The test item falls within the applicability domain of the model and was therefore reliably predicted for its acute toxicity (48h-EC50) to daphnids. Therefore, this endpoint value can be considered valid for use in risk assessment and classification and labelling.

The 48h-EC50 of the test item to daphnids was predicted as 1.2 mg/L (95% CI: 0.84 - 1.6 mg/L).