4-methyl-2-(pentan-3-yl)tetrahydro-2H-pyran-4-ol

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

2009-07-24 to 2009-09-04

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

This study was performed according to OECD Guideline 201 with GLP certificate. All validity criteria were fulfilled. However, this study is considered reliable with restrictions due to missing information on isomeric composition of the test substance and the use of solvent. Indeed, 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 sufficiently high water solubility of the substance (6,89 g/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 0.5 mL/L, corresponding to 395 mg/L (with a density of 0.79), which is 5 times higher than the recommended maximum level of solvent (below 0.1 mL/L; OECD No. 23) but is below the NOEC of acetone (which was reported in the ECHA disseminated dossier at 530 mg/L). Finally, the % inhibition of growth rate for the limit test for each day were not calculated in the study report. No calculation details was reported for the 7% inhibition after 72 hours. The result was based on 100 mg/L while the nominal concentration was 98,8 mg/L and the measured concentrations 105 mg/L.

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

not specified

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Remarks:

Inspection on July 15-16 2008, implementation on July 16 2009.

Specific details on test material used for the study:

- Storage condition of test material: Store at room temperature and keep away from direct sun light.

Analytical monitoring:

yes

Remarks:

The test item application rate and the stability of the test substance in water throughout the test period were checked by appropriate analytical verification of the test solutions of the limit test at both test initiation and test completion.

Details on sampling:

- Concentrations: Range finding test (nominal concentrations): 0.01 - 0.1 - 1.0 - 9.9 and 98.7 mg/L ; limit test : 98.8 mg/L
- Sampling method: The test item was quantified in the treatment solutions used for the limit test at both test initiation and test completion, in one replicate unit at random, according to previously validated method.

Vehicle:

yes

Remarks:

acetone

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The treatment solutions were prepared in acetone. The concentrations of the treatment solutions were set in such a manner that treatment concentrations were achieved when delivering 50 µL of the treatment solutions per test vessel. For the reference item, a stock solution was prepared using 0.043 g of potassium dichromate in 10 mL of water. The stock solution was further diluted 1/10 v./v in water. Aliquots of 0.050, 0.150, 0.185, 0.250 mL were added to the test vessels and the volumes adjusted with reconstituted water such to achieve treatment concontrations of 0.2, 0.6, 0.75 and 1.0 mg/L.

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Source (laboratory, culture collection): The strain was provided by in December 2007 by the Muséum National d'Histoire Naturelle (Paris, France) and regularly sub-cultured in the OECD medium at Phytosafe site.
- Age of inoculum (at test initiation): The inoculum culture was prepared 2-4 days before the start of the test and incubated under the same conditions as the test cultures such to adapt the test algae to test conditions and ensure that the algae were in the exponential growth phase when used to inoculate th etest solutions.

ACCLIMATION
- Culturing media and conditions (same as test or not): Same as test.

Test type:

static

Water media type:

other: reconstituted water

Limit test:

yes

Total exposure duration:

72 h

Remarks on exposure duration:

None

Post exposure observation period:

None

Hardness:

Not reported

Test temperature:

21 - 24 °C

pH:

8.4-8.65 at t0
8.27-8.31 at t72h

Dissolved oxygen:

Not reported

Salinity:

Not applicable

Nominal and measured concentrations:

A range finding test was performed, testing the following nominal concentrations: 0.01 - 0.1 - 1.0 - 9.9 and 98.7 mg/L.
A limit test was performed at a concentration of 98.8 mg/L.

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass Erlenmeyer flasks (250mL) filled with 100 mL of culture served as test vessels. They were capped with air-permeable stoppers.
- Initial cells density: 2 to 5 x 10 E-3 cells/mL
- No. of vessels per concentration (replicates): 6 replicate units for the 100 mg/L treated groups, 3 replicate units for each of four concentrations of Potassium dichromate
- No. of vessels per control (replicates): 6 replicate units for the controls (water control, solvent control)

GROWTH MEDIUM
- Standard medium used: yes, OECD medium from OECD TG 201 according to ISO 8692


OTHER TEST CONDITIONS
- Light intensity and quality: Cultures received continuous uniform fluorescent illumination within 4440 - 8880 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: electronic cell counter (Coulter Counter ZM)

TEST CONCENTRATIONS
- Spacing factor for test concentrations:
- Justification for using less concentrations than requested by guideline:
- Range finding study : yes, testing the following cocnentrations : 0.01 - 0.1 - 1.0 - 9.9 and 98.7 mg/L.
- Test concentrations: one limit concentration tested of 98.8 mg/L.
- Results used to determine the conditions for the definitive study: In the range-finding test, neither the specific growth rate nor the yield was affected in the 100 mg/L test item treated unit. The definitive test was thus performed as a limit test for the 100 mg/L treatment concentration.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate was used as reference substance.

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

- Exponential growth in the control (for algal test): yes, the multiplication rate was approximately 25.

Results with reference substance (positive control):

- Results with reference substance valid?
- EC50 72h (specific growth rate)= 0.76 mg/L

Reported statistics and error estimates:

NOEC was derived from the F-variance analysis at a 5%-confidence level of the response variable (specific growth rate) in each treated group as compared to the control group.
An additional T.test was performed in order to re-evaluate the results of the study report.

Table 2 : Analysis of the test solutions

The test material was quantified in the treatment solutions used for the limit test at both test initiation and test completion, in one replicate unit taken at random.

Nominal (mg/L)	Measured concentration (mg/L)	Percent recovery (%)
Test initiation	103.3	104.6
	104.6	105.9
Mean	-	105.2
Test completion	104.2	105.5
	104.0	105.3
Mean	-	105.4

Table 3 : Measured % inhibition of specific growth rate per day (section-by section and total), for the range-finding test and the limit test based on the water control

Time \ Treatment (mg/L)	0,01  (RF test)	0,1  (RF test)	1  (RF test)	9,9  (RF test)	98,7  (RF test)	98,8 (limit test) *
0 to 24 h	-6,00	-25,60	-19,90	12,30	-7,80	-4,76*
24 to 48 h	-19,00	-4,00	21,20	-18,40	34,70	-5*
48 to 72 h	h-48,90	-42,60	-72,30	31,50	-41,50	6,74*
total period	-22,30	-22,30	-19,00	6,30	-1,30	-0,65*

* The % inhibition of growth rate for the limit test (98.8 mg/L) for each day were calculated by the study reviewer, taking the water control of the limit test into account. These results were not mentionned in the study report. On the last day of testing (48 to 72 h), it was indicated in the study report that the % inhibition was 7.1 % (no calculation details). After calculation by the reviewer, 6.74 % was obtained. In contradiction with the study report, this result was not found to be significant after performing an additional T.test (see Table 5 in Overall remarks).

Moreover, the % inhibition of specific growth rate were re-calculated taking the solvent control into account (see Table 6 in Overall remarks).

Table 4 : Measured specific growth rates per day (section-by-section and total), for the range-finding test (RF test) and the limit test

Time \ Treatment (mg/L)	solvent control (limit test)	water control (limit test)	solvent control (RF test)	water control  (RF test)	0,01  (RF test)	0,1  (RF test)	1  (RF test)	9,9  (RF test)	98,7  (RF test)	98,8 (limit test)
0 à 24	1,48	1,47	1,23	1,19	1,26	1,50	1,43	1,05	1,28	1,54
24 à 48	1,46	1,40	1,27	1,18	1,41	1,23	0,93	1,40	0,77	1,47
48 à 72	1,71	1,78	1,05	0,87	1,30	1,24	1,50	0,60	1,23	1,66
total period	1,55	1,55	1,18	1,08	1,32	1,32	1,29	1,02	1,10	1,56

The NOEC obtained for growth rate is > 100 mg/L.

The ErC50 based on specific growth rate is > 100 mg/L.

Validity criteria fulfilled:

yes

Conclusions:

The over-all value for the 72-hour testing period was considered as similar to that of the control. The NOEC for growth rate was > 100 mg/L and the ErC50 based on specific growth rate was > 100 mg/L.
Even if all validity criteria were fulfilled, this study is considered reliable with restrictions.

Executive summary:

The influence of the test substance on algal growth inhibition (Desmodesmus subspicatus) was investigated according to OECD Guideline 201. Algae were exposed to the test substance under static conditions during 72 hours.

Based on the results of a range-finding test, a limit test was performed at a concentration of 98.8 mg/L. Six replicate units were performed for the solvent control, the water control and the 98.8 mg/L treated group (limit test). The test item concentration was quantified in the treatment solutions used for the limit test at both test initiation and completion, in one replicate unit taken at random. The resulting solution was assessed by GC/FID, using external calibration.

The over-all value for the 72-hour testing period was considered as similar to that of the control. The NOEC for growth rate was > 100 mg/L and the ErC50 based on specific growth rate was > 100 mg/L.

Even if all validity criteria were fulfilled, this study is considered reliable with restrictions for the following reasons:

- The substance is adequately identified, but some data on composition is missing.

- 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. The use of solvent is not necessary or even useful to test soluble test substances.

- Considering the sufficiently high water solubility of the substance (6,89 g/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 0.5 mL/L, corresponding to 395 mg/L (with a density of 0.79), which is 5 times higher than the recommended maximum level of solvent (below 0.1 mL/L; OECD No. 23) but is below the NOEC of acetone (which was reported in the ECHA disseminated dossier at 530 mg/L).

- The % inhibition of growth rate for the limit test for each day were not calculated in the study report. No calculation details was reported for the 7% inhibition after 72 hours.

- The result was based on 100 mg/L while the nominal concentration was 98,8 mg/L and the measured concentrations 105 mg/L.

Reference 2

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

(Q)SAR

Adequacy of study:

key study

Study period:

October 02nd, 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® toolbox – in Silico Algorithms For Environmental Risk And Toxicity version 2.4

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

3. SMILES OR OTHER IDENTIFIERS USED AS INPUT FOR THE MODEL
CC1(O)CC(OCC1)C(CC)CC
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 = 6886 mg/L (or -1.432 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

Reason / purpose for cross-reference:

reference to other study

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition 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 toxicity to algae as would be expected in a laboratory experiment following the OECD Guideline 201 and EC method C.3 for specific, named mechanisms of action. The model provides an in silico prediction for the 72-hour ErC50 value that can effectively be used in place of an experimentally derived 72-hour ErC50 value. The regression is method used to achieve this has been fully validated following the OECD (2004) recommendations.

GLP compliance:

no

Specific details on test material used for the study:

- log KOW = 2.20 (Phytosafe, 2009; EU Method A.8, Shake Flask Method)
- Water solubility: 6886 mg/L at 25°C (KREATiS, 2019)
- Mechanism of action: MechoA 1.1: non-polar narcosis (Bauer et al., 2018)

Analytical monitoring:

no

Details on sampling:

not applicable

Vehicle:

no

Details on test solutions:

not applicable

Test organisms (species):

other: Pseudokirchneriella subcapitata, Desmodesmus subspicatus, Scenedesmus quadricauda

Details on test organisms:

No difference in terms of toxic mechanism of action between algae (or indeed other) aquatic species is expected. Any observed differences may be attributed to lifestyle related parameters and relative duration of study versus cell size 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:

72 h

Remarks on exposure duration:

Results from a test duration of 72 hours only were used for this algorithm.

Post exposure observation period:

None

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 25 °C depending on the species used to construct the algorithm. This small difference is not expected to contribute to the variability of the ErC50 values found in experimental data.

pH:

Test results were preferably taken from studies with measured pHs between 6 - 9. However it is recognized that in some cases (due to high luminosity) the pH may increase in the control and lower concentrations (which do not cause significant effect over the study period). This pH increase did not generally disqualify the study from being used in the test and validation set for non-polar chemicals.

Dissolved oxygen:

The QSAR is based on data from reliable studies performed at acceptable oxygen concentrations.

Salinity:

Not applicable

Conductivity:

No data

Nominal and measured concentrations:

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

Details on test conditions:

Following the guideline OECD 201, all studies were from a static test design. 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:

72 h

Dose descriptor:

EC50

Effect conc.:

145 mg/L

Nominal / measured:

meas. (not specified)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95%CL: 124 - 168 mg/L

Details on results:

The test item falls within the applicability domain of the model and was therefore reliably predicted for its toxicity (72h-ErC50) to algae. Therefore, this endpoint value can be considered valid for use in risk assessment and classification and labelling.

Results with reference substance (positive control):

Not applicable

Reported statistics and error estimates:

95% confidence interval (α = 0.05): 124 – 168 mg/L
QSAR statistical parameters are given in the QMRF and the QPRF

Analysis of the Applicability Domain of the model

Descriptor domain

The Subcooled Liquid Water Solubility value (-1.432 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 -4.38 to 0.49.

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 a simple hydroxylated ether is expected to exert a MechoA 1.1 and can be taken into account by the model.

Validity criteria fulfilled:

yes

Remarks:

fully

Conclusions:

The test item falls within the applicability domain of the model and was therefore reliably predicted for its toxicity (72h-ErC50) to algae. Therefore, this endpoint value can be considered valid for use in risk assessment and classification and labelling.

The 72h-ErC50 of the test item to algae was predicted as 145 mg/L.
95% confidence interval (α = 0.05): 124 – 168 mg/L

Executive summary:

A Quantitative Structure-Activity Relationship (QSAR) was used to calculate the inhibition of growth to algae of the test item. This 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 Guideline for Testing of Chemicals No. 201, "Freshwater Alga and Cyanobacteria, Growth Inhibition Test" (OECD, 2006), referenced as Method C.3 of Commission Regulation No. 440/2008 (European Commission, 2008). The criterion predicted was the ErC50 (Median Effective Concentration for specific growth rate), a statistically derived concentration which is expected to cause 50% inhibition of intrinsic rate of growth of the test system within a period of 72 hours.

The growth inhibition of algae was determined using validated QSAR model for the Mechanism of Action (MechoA) in question (MechoA 1.1, i.e. non-polar narcosis) (Bauer et al., 2018). The QSAR model is based on validated data for a training set of 40 chemicals derived from 72-hour ErC50 test on algae, for which the concentrations of the test item had been determined by chemical analyses over the test period.

The test item falls within the applicability domain of the model and was therefore reliably predicted for its toxicity (72h-ErC50) to algae. Therefore, this endpoint value can be considered valid for use in risk assessment and classification and labelling.

The 72h-ErC₅₀of the test item to algae was predicted as 145 mg/L.

95% confidence interval (α = 0.05): 124 – 168 mg/L

Description of key information

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

72h-ErC50 Algae = 145 mg/L (95% confidence interval: 124 – 168 mg/L)

Key value for chemical safety assessment

EC50 for freshwater algae:

145 mg/L

Additional information

One experimental study and one QSAR prediction are available to assess the toxicity of the registered substance to aquatic algae.

The experimental study (Phytosafe, 2009) was performed according to OECD Guideline 201 with GLP statement but was considered reliable with restrictions due to missing information on the isomeric composition of the test substance and the use of solvent. Indeed, 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 necessary for water soluble substances. Considering the high water solubility of the substance (6,89 g/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 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. Furthermore, the concentration/quantity of solvent used in the treatment solutions was 0.5 mL/L, corresponding to 395 mg/L (with a density of 0.79), which is 5 times higher than the recommended maximum level of solvent (below 0.1 mL/L; OECD No. 23) but is below the NOEC of acetone (which was reported in the ECHA disseminated dossier at 530 mg/L).

Under the test conditions, the 72h-ErC50 and NOEC values reported in the study report, based on growth rate of the algae Desmodesmus subspicatus, were greater than 100 mg/L, the limit test concentration. The result was based on 100 mg/L while the nominal concentration was 98,8 mg/L and the measured concentrations was 105 mg/L. However, this result supports the (more quantitative) key data presented below.

The QSAR prediction (KREATiS, 2019) was considered as reliable and was used as key data. 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 201. The growth inhibition of algae was determined using validated QSAR model for the Mechanism of Action (MechoA) in question (MechoA 1.1, i.e. non-polar narcosis) (Bauer et al., 2018). The QSAR model is based on validated data for a training set of 40 chemicals derived from 72-hour ErC50 test on algae, for which the concentrations of the test item had been determined by chemical analyses over the test period. The result below is the toxicity values anticipated during a 72-hour study on algae based on measured concentrations. Therefore, this endpoint value can be considered valid for use in risk assessment and classification and labelling.

The 72h-ErC50 of the test item to algae was predicted as 145 mg/L (95% CI: 124– 168 mg/L)

This quantitative value was considered as the key value for chemical safety assessment.