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Toxicity to aquatic algae and cyanobacteria

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Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Study period:
not reported
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
other: Read-across from an analogue substance for which there is available information (Klimish = 4).
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
See cross-reference to justification of read-across.
Reason / purpose:
read-across source
Reason / purpose:
read-across: supporting information
Specific details on test material used for the study:
¡
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 1.6 mg/L
Nominal / measured:
not specified
Conc. based on:
not specified
Basis for effect:
growth rate
Remarks on result:
other: Based on the read-across from an analogue substance.
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
1.6 mg/L
Nominal / measured:
not specified
Conc. based on:
not specified
Basis for effect:
growth rate
Remarks on result:
other: Based on the read-across from an analogue substance.
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 21 mg/L
Nominal / measured:
not specified
Conc. based on:
not specified
Basis for effect:
cell number
Remarks on result:
other: Based on the read-across from an analogue substance.
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
> 21 mg/L
Nominal / measured:
not specified
Conc. based on:
not specified
Basis for effect:
cell number
Remarks on result:
other: Based on the read-across from an analogue substance.
Conclusions:
Based on the read-across from an analogue substance, the 72h-NOEC and the 72h-EC50 was 1.6 and > 1.6 mg/L, respectively based on growth rate and 21 and > 21 mg/L based on the cell number.
Executive summary:

Based on the read-across from an analogue substance, the 72h-NOEC and the 72h-EC50 was 1.6 and > 1.6 mg/L, respectively based on growth rate and 21 and > 21 mg/L based on the cell number.

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
(Q)SAR
Adequacy of study:
weight of evidence
Study period:
not applicable (calculated data)
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Calculation with a valid QSAR (ECOSAR), l-limonene falls within the applicability domain of the (Q)SAR model, the prediction fits for the regulatory purpose, and the information is enough documented.
Justification for type of information:
1. Relevance of the model

ECOSAR v1.00 is reported as a usable valid model by ECHA (R6: QSARs and grouping of chemicals, May 2008).
ECOSAR contains a library of class-based QSARs for predicting aquatic toxicity, overlaid with an expert decision tree for selecting the appropriate chemical class. ECOSAR Version 1.00 is programmed to identify over 120 chemical classes and allows access to over 600 QSARs for numerous endpoints and organisms.
The structure-activity relationships (SARs) presented in ECOSAR v1.00 are used to predict the aquatic toxicity of chemicals based on their similarity of structure to chemicals for which the aquatic toxicity has been previously measured. SARs are developed for chemical classes based on measured test data that have been submitted by industry to the US-Environmental Protection Agency or collected from publicly available sources. To date, over 440 SARs have been developed for more than 120 chemical classes. The supporting data sets (training sets) used to derive SARs within a chemical class range from the very large, e.g., neutral organics, to the very small, e.g., aromatic diazoniums. The class with the greatest number of SARs based on measured data is the neutral organics class, which has SARs ranging from acute and chronic toxicity to fish to a 14-day LC50 SAR for earthworms in artificial soil.
The ECOSAR Class Program is a computerized version of the ecotoxicity analysis procedures as currently practiced by the Office of Pollution Prevention and Toxics (OPPT) when data are lacking for regulatory endpoints. It has been developed within the regulatory constraints of the Toxic Substances Control Act (TSCA). It is a pragmatic approach to SAR as opposed to a theoretical approach.

2. Validation of the model

- Endpoint (OECD Principle 1)
Th endpoints are the short term (96h-EC50) and the chronic effects of the substance on the exponential growth rate of the algae expressed in mg/L. The ChV, or Chronic Value, is defined as the geometric mean of the no observed effect concentration (NOEC) and the lowest observed effect concentration (LOEC). This can be mathematically represented as: ChV = 10^([log (LOEC x NOEC)]/2).
The studies collected for the training set chemicals in ECOSAR undergo an extensive data validation step to ensure appropriateness for inclusion in the model. For freshwater algae, species frequently include Desmodesmus subspicatus or Pseudokirchneriella subcapitata. ECOSAR study criteria articulate that the toxicity should be measured at pH 7 (replicating environmental conditions), the total organic carbon content should not exceed 2 mg/L, the water hardness should be approximately 150 mg/L CaCO3, results should be adjusted to, or measured at, 100% active ingredient, and flow-through measured is preferred over static nominal, etc. Data received or identified in the open literature which is not accompanied with full study details to confirm conditions are often not considered appropriate for model development.

- Algorithm (OECD Principle 2)

Acute toxicity:
The green algae 96-h EC50 values used to develop this SAR were measured using the computer program, KOWWIN (Version 1.67). The SAR equation used to estimate toxicity is: Log 96-h EC50 (mmol/L) = -0.6271 log Kow + 0.5687
 The EC50 is in millimoles per liter (mM/L); To convert the EC50 from mM/L to mg/L, multiply by the molecular weight of the compound.

Chronic toxicity
The green algae chronic values (ChV) used to develop this SAR were measured and the octanol water partition coefficients (Kow) were calculated using the computer program, KOWWIN (Version 1.67). The SAR equation used to estimate toxicity is:
 Log ChV (mmol/L) = -0.5547 log Kow + 0.0061
The ChV is in millimoles per liter (mM/L); To convert the ChV from mM/L to mg/L, multiply by the molecular weight of the compound.


-Applicability domain (OECD Principle 3)
Neutral organic chemicals are non ionizable and non reactive and act via simple nonpolar narcosis generally thought of as a reversible, drug-induced loss of conscience (general anesthesia). This general narcosis is often referred to as baseline toxicity (Franks and Lieb 1990, Veith and Broderius 1990). Application: Solvents, non-reactive, non-ionizable neutral organic compounds:
1. Alcohols
2. Acetals
3. Ketones
4. Ethers
5. Alkyl halides
6. Aryl halides
7. Aromatic hydrocarbons
8. Halogenated aromatic hydrocarbons
9. Halogenated aliphatic hydrocarbons
10. Sulfides and di-sulfides

ECOSAR v.1.00 SAR Limitations:
Maximum Kow: 6.4 (96h-EC50) and 8 (ChV)
Maximum MW: 1000

- Uncertainty of the prediction (OECD Principle 4)
Acute toxicity: N = 51 + 11; and the Coefficient of Determination (R2) = 0.5956.
Chronic toxicity: N = 43 + 7; and the Coefficient of Determination (R2) = 0.7187.

3. Adequacy of result for classification & labelling an/or risk assessment
According to the main criteria used by Hulzebos and Posthumus, ECOSAR is a reliable QSAR to predict 96h-EC50 for algae. The ECOSAR prediction of toxicity of l-limonene on invertebrates is adequate for the purpose of risk assessment and classification and labelling.
Qualifier:
according to
Guideline:
other: ECHA guidance on QSARs. R6
Deviations:
no
Principles of method if other than guideline:
ECOSAR model for neutral organics.


GLP compliance:
no
Remarks:
The data were calculated by a computer model
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
Not applicable (calculated data)
Analytical monitoring:
no
Details on sampling:
Not applicable (calculated data)
Vehicle:
no
Details on test solutions:
Not applicable (calculated data)
Test organisms (species):
other: For freshwater algae, species frequently include Desmodesmus subspicatus or Pseudokirchneriella subcapitata.
Details on test organisms:
Not applicable (calculated data)
Test type:
other: flow-through measured is preferred over static nominal
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h
Post exposure observation period:
Not applicable (calculated data)
Hardness:
ECOSAR study criteria articulate that the water hardness should be approximately 150 mg/L CaCO3
Test temperature:
Not applicable (calculated data)
pH:
ECOSAR study criteria articulate that the toxicity should be measured at pH 7 (replicating environmental conditions)
Dissolved oxygen:
Not applicable (calculated data)
Salinity:
Not applicable
Nominal and measured concentrations:
Not applicable (calculated data)
Details on test conditions:
Not applicable (calculated data)
Reference substance (positive control):
not required
Key result
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
0.904 mg/L
Nominal / measured:
estimated
Conc. based on:
test mat.
Basis for effect:
growth rate
Key result
Duration:
96 h
Dose descriptor:
other: Geometric mean of NOEC and LOEC
Effect conc.:
0.514 mg/L
Nominal / measured:
estimated
Conc. based on:
test mat.
Basis for effect:
growth rate
Details on results:
1. Relevance of the model

ECOSAR v1.00 is reported as a usable valid model by ECHA (R6: QSARs and grouping of chemicals, May 2008).
ECOSAR contains a library of class-based QSARs for predicting aquatic toxicity, overlaid with an expert decision tree for selecting the appropriate chemical class. ECOSAR Version 1.00 is programmed to identify over 120 chemical classes and allows access to over 600 QSARs for numerous endpoints and organisms.
The structure-activity relationships (SARs) presented in ECOSAR v1.00 are used to predict the aquatic toxicity of chemicals based on their similarity of structure to chemicals for which the aquatic toxicity has been previously measured. SARs are developed for chemical classes based on measured test data that have been submitted by industry to the US-Environmental Protection Agency or collected from publicly available sources. To date, over 440 SARs have been developed for more than 120 chemical classes. The supporting data sets (training sets) used to derive SARs within a chemical class range from the very large, e.g., neutral organics, to the very small, e.g., aromatic diazoniums. The class with the greatest number of SARs based on measured data is the neutral organics class, which has SARs ranging from acute and chronic toxicity to fish to a 14-day LC50 SAR for earthworms in artificial soil.
The ECOSAR Class Program is a computerized version of the ecotoxicity analysis procedures as currently practiced by the Office of Pollution Prevention and Toxics (OPPT) when data are lacking for regulatory endpoints. It has been developed within the regulatory constraints of the Toxic Substances Control Act (TSCA). It is a pragmatic approach to SAR as opposed to a theoretical approach.

2. Validation of the model

- Endpoint (OECD Principle 1)
Th endpoints are the short term (96h-EC50) and the chronic effects of the substance on the exponential growth rate of the algae expressed in mg/L. The ChV, or Chronic Value, is defined as the geometric mean of the no observed effect concentration (NOEC) and the lowest observed effect concentration (LOEC). This can be mathematically represented as: ChV = 10^([log (LOEC x NOEC)]/2).
The studies collected for the training set chemicals in ECOSAR undergo an extensive data validation step to ensure appropriateness for inclusion in the model. For freshwater algae, species frequently include Desmodesmus subspicatus or Pseudokirchneriella subcapitata. ECOSAR study criteria articulate that the toxicity should be measured at pH 7 (replicating environmental conditions), the total organic carbon content should not exceed 2 mg/L, the water hardness should be approximately 150 mg/L CaCO3, results should be adjusted to, or measured at, 100% active ingredient, and flow-through measured is preferred over static nominal, etc. Data received or identified in the open literature which is not accompanied with full study details to confirm conditions are often not considered appropriate for model development.

- Algorithm (OECD Principle 2)

Acute toxicity:
The green algae 96-h EC50 values used to develop this SAR were measured using the computer program, KOWWIN (Version 1.67). The SAR equation used to estimate toxicity is: Log 96-h EC50 (mmol/L) = -0.6271 log Kow + 0.5687
 The EC50 is in millimoles per liter (mM/L); To convert the EC50 from mM/L to mg/L, multiply by the molecular weight of the compound.

Chronic toxicity
The green algae chronic values (ChV) used to develop this SAR were measured and the octanol water partition coefficients (Kow) were calculated using the computer program, KOWWIN (Version 1.67). The SAR equation used to estimate toxicity is:
 Log ChV (mmol/L) = -0.5547 log Kow + 0.0061
The ChV is in millimoles per liter (mM/L); To convert the ChV from mM/L to mg/L, multiply by the molecular weight of the compound.


-Applicability domain (OECD Principle 3)
Neutral organic chemicals are non ionizable and non reactive and act via simple nonpolar narcosis generally thought of as a reversible, drug-induced loss of conscience (general anesthesia). This general narcosis is often referred to as baseline toxicity (Franks and Lieb 1990, Veith and Broderius 1990). Application: Solvents, non-reactive, non-ionizable neutral organic compounds:
1. Alcohols
2. Acetals
3. Ketones
4. Ethers
5. Alkyl halides
6. Aryl halides
7. Aromatic hydrocarbons
8. Halogenated aromatic hydrocarbons
9. Halogenated aliphatic hydrocarbons
10. Sulfides and di-sulfides

ECOSAR v.1.00 SAR Limitations:
Maximum Kow: 6.4 (96h-EC50) and 8 (ChV)
Maximum MW: 1000

- Uncertainty of the prediction (OECD Principle 4)
Acute toxicity: N = 51 + 11; and the Coefficient of Determination (R2) = 0.5956.
Chronic toxicity: N = 43 + 7; and the Coefficient of Determination (R2) = 0.7187.

3. Adequacy of result for classification & labelling an/or risk assessment
According to the main criteria used by Hulzebos and Posthumus, ECOSAR is a reliable QSAR to predict 96h-EC50 for algae. The ECOSAR prediction of toxicity of l-limonene on invertebrates is adequate for the purpose of risk assessment and classification and labelling.
Results with reference substance (positive control):
Not applicable (calculated data)
Reported statistics and error estimates:
Not applicable (calculated data)

No data

Validity criteria fulfilled:
yes
Remarks:
Log Kow < 5; MW = 136 g/mol
Conclusions:
The ECOSAR model to estimate acute and chronic toxicity in a 96 hours freshwater algae study estimated the EC50 and the NOEC were 0.904 mg/L and 0.514 mg/L, respectively.
Executive summary:

The acute toxicity of l-limonene to aquatic invertebrates was estimated using the ECOSAR model from EPISUITE 4.1 reported as a valid model by ECHA (R6: QSAR and grouping of chemicals, May 2008). ECOSAR computes toxicity based on a linear mathematical relationship between the predicted log Kow values and the corresponding log of the measured toxicity values (mmol/L). l-limonene feld in the applicability domain of this QSAR model. The calculation was ran using the experimental log Kow value of 4.38 and water solubility of 12.3 mg/L for d-limonene asthese values are expected to besimilar for bothenantiomers.

Results of computer modelling to estimate acute and chronic toxicity in a 96 -hour freshwater algae study show that the EC50 and the NOEC were 0.904 mg/L and 0.514 mg/L, respectively.

This ECOSAR prediction of acute and chronic toxicity of l-limonene on algae is adequate for the purpose of risk assessment and classification and labelling.

Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Read-across from an analogue substance for which there is available information (Klimish=2).
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
See cross-reference to justification of read-across.
Reason / purpose:
read-across source
Reason / purpose:
read-across: supporting information
Duration:
72 h
Dose descriptor:
NOELR
Effect conc.:
50 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Remarks:
WAF
Basis for effect:
growth rate
Remarks on result:
other: Interpretation of data, Report states wrong value
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
2.62 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
dissolved
Remarks:
IR (Hydrocarbons)
Basis for effect:
growth rate
Remarks on result:
other: Interpretation of data, Report gives wrong values
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
150 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Remarks:
WAF
Basis for effect:
growth rate
Remarks on result:
other: Interpretation of data; deviates from report
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
ca. 8 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
dissolved
Remarks:
IR (hydrocarbons)
Basis for effect:
growth rate
Remarks on result:
other: Interpretation of data; deviates from report
Conclusions:
Based on the read-across from an analogue substance the 72h-NOELr and 72h-ELr50 towards Desmodesmus subspicatus were estimated to be 50 and 150 mg/l, respectively. The 72h-NOECr and 72h-ECr50 were circa 8 and 2.6 mg/l (measured, hydrocarbons).
Executive summary:

Based on the read-across from an analogue substance the 72h-NOELr and 72h-ELr50 towards Desmodesmus subspicatus were estimated to be 50 and 150 mg/l, respectively. The 72h-NOECr and 72h-ECr50 were circa 8 and 2.6 mg/l (measured, hydrocarbons).

Description of key information

Weight of evidence: A study performed according to OECD guideline 201 under GLP investigated the toxicity of orange oil (containing typically >95% of d-limonene) towards Desmodesmus subspicatus. The result is comparable to the result of a non assignable MITI study performed on Pseudokircheneriella subcapitata with dipentene, the racemic of limonene. These studies suggest that both enantiomers have the same level of toxicity on algae. However, due to the uncertainities on the purity of the substances tested, the slight lower toxicity values calculated for l-limonene with a valid ECOSAR model are considered as key values for the purpose of risk assessment and classification&labelling.

Key value for chemical safety assessment

EC50 for freshwater algae:
0.904 mg/L
EC10 or NOEC for freshwater algae:
0.514 mg/L

Additional information

Reliable experimental values for algae, a 72h-ErC50 and a 72h-NOErC of 8 and 2.62 mg/L (measured hydrocarbons), respectively, were derived from a study conducted with orange oil on Desmodesmus subspicatus using WAFs. Although the composition of orange oil was not reported, these values were considered relevant, as typical content of d-limonene in orange oil is > 95% and this study was performed according to OECD Guideline under GLP.

These values are comparable with the result of a MITI study performed by the Japanese authority on Pseudokircheneriella subcapitata with dipentene, the racemic form of limonene, giving a 72h-ErC50 and a 72h-NOErC of > 1.6 and 1.6 mg/L, respectively. This study was assessed as non assignable because it is poorly documented however it is considered as a reliable source and it suggests that both enantiomers have the same level of toxicity on algae.

Finally, the software ECOSAR, a valid QSAR, was used to assess the toxicity of l-limonene. The result was calculated with its measured Kow value and with the measured water solubility value of d-limonene which is expected to be similar for both enantiomers. The estimated values for algae were lower than the experimental values with a 96h-EC50 of 0.904 mg/L and a chronic toxicity value of 0.514 mg/L. The ECOSAR model has been validated to estimate the toxicity of l-limonene on algae.

Therefore, the estimated values by the ECOSAR model are considered as the reliable values for the purpose of risk assessment and classification&labelling.