Schinus terebinthifolius, ext.

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:

From 2017-06-28 to 2017-06-29

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

Remarks:

This calculation method 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", referenced as Method C.3 of Commission Regulation No. 440/2008. This approach uses a thermodynamically based module to calculate the water concentrations and the individual toxicity of each constituent is first derived from a QSAR model validated to be compliant with the OECD recommandations for QSAR modeling (OECD, 2004). Then, the effective loading rate of the mixture to the test organisms (i.e. Pseudokirchneriella subcapitata) is determined as a Water Accommodated Fraction (WAF) test (OECD, 2000).

Justification for type of information:

1. SOFTWARE
iSafeRat® HA-QSAR toolbox v2.3

2. MODEL (incl. version number)
iSafeRat® holistic HA-QSAR v1.7

3. SMILES OR OTHER IDENTIFIERS USED AS INPUT FOR THE MODEL
See attached report

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

5. APPLICABILITY DOMAIN
See attached report

6. ADEQUACY OF THE RESULT
See attached report

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:

This calculation method predicts the endpoint value which would be expected when testing the substance under experimental conditions in a laboratory following OECD Guideline 202 adapted for testing as a mixture using the WAF method. The criterion predicted was the effective loading rate of the mixture EL50 (Median Effective Loading), a statistically derived loading rate which is expected to cause immobility in 50% of test animals within a period of 48 hours.
This approach uses a thermodynamically based module to calculate the water concentrations and the individual toxicity of the main constituents are first derived from a QSAR model validated to be compliant with the OECD recommendations for QSAR modelling (OECD, 2004). Then the effective loading rate of the mixture to the test organisms (i.e. Daphnia magna) is determined as a Water Accommodated Fraction (WAF) test (OECD, 2000).

GLP compliance:

no

Specific details on test material used for the study:

No additional information

Analytical monitoring:

no

Details on sampling:

Not applicable

Vehicle:

no

Details on test solutions:

Not applicable

Test organisms (species):

Daphnia sp.

Details on test organisms:

Not applicable

Test type:

other: QSAR

Water media type:

freshwater

Limit test:

no

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

Reference substance (positive control):

no

Remarks:

QSAR model

Key result

Duration:

48 h

Dose descriptor:

EL50

Effect conc.:

5.1 mg/L

Nominal / measured:

estimated

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

The 48h-EL50 is predicted at 5.1 mg/L. At this loading rate, the expected concentrations of each constituent in the mixture (based on thermodynamic calculation) are specify in the attached report.
This method has previously been validated in an internal publication (Bicherel & Thomas, 2014, available in "Attached background material") and will be made available to the Competent Authorities upon request.

Results with reference substance (positive control):

Not applicable

Reported statistics and error estimates:

Not applicable

No additional information

Validity criteria fulfilled:

not applicable

Conclusions:

The 48-h EL50 based on mobility was determined to be 5.1 mg/L.

Executive summary:

A calculation method prediction was performed to assess the acute toxicity to Daphnia magna of the test substance, a Natural Complex Substance, using the WAF method. This calculation method 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" (1), referenced as Method C.2 of Commission Regulation No. 440/2008 (2) adapted for testing of a mixture using the WAF method. The criterion predicted was the median effective loading rate of the mixture EL50 (Median Effect Loading), a statistically derived loading rate which is expected to cause immobility in 50% of test animals within a period of 48 hours.

 

The acute toxicity to the daphnids was determined using a calculation method for the Mode of Action (MOA) in question (MOA 1, non-polar narcosis). This method has previously been validated in an internal publication (Bicherel and Thomas, 2014). This algorithm is based on a QSAR model which has been validated to be compliant

with the OECD recommandations for QSAR modeling (OECD, 2004). The QSAR model is based on validated data for a training set of 58 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. Further to this the effective loading rate of the WAF is determined by using a series of calculation steps using phase equilibrium thermodynamics and excluding the non-bioavailable fraction, this approach is based on validated data derived from 48-hour EL50 tests on daphnid, for which the concentrations of the test item had been determined by chemical analyses over the test period.

The 48-h EL50 based on mobility was determined to be 5.1 mg/L.

Reference 2

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

10-14 November 2008

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

This GLP study was performed according to OECD Guideline 202 with the following deficiencies: old WAF method - dispersions were stirred for 3 hours and then filtered ; the 48-hour EL50 was determined as the geometric mean value of the two consecutive test concentrations with 0 and 100% immobility (EL0 and EL100) as no dose/response curve were observed after 48 hours.

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)

Deviations:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Remarks:

The Swiss GLP Monitoring Authorities (inspected on 05-09 & 26-30 November 2007 / signed on 12 November 2008)

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Appearance: Pale yellow liquid

Analytical monitoring:

yes

Details on sampling:

Analysis of the test Item concentrations
For the characterization of the WAFs, duplicate samples were taken from each treatment before the test start and at the end of the test after 48 hours.
For the 48-hour stability samples, the contents of the respective replicates were combined prior to sampling.
Immediately after sampling, acetone was added to each sample (1% of acetone per sample volume). All samples as listed below were analyzed immediately after sampling.
The concentrations of the test item, based on the sum of four peaks, were analyzed in the duplicate test media samples of all loading rates from both sampling times (0 and 48 hours).
From the control, only one of the duplicate samples was analyzed from each of the sampling times.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: For preparation of the WAFs, individual dispersions of the test item with the loading rates as mentioned above were prepared (table 6.1.3/1)
The dispersions were stirred for 3 hours to dissolve a maximum amount of the different compounds of the test item in the dispersion. Then, the dispersions were filtered through membrane filters (Schleicher & Schuell, Type NC45, pore size 0.45 μm) and the undiluted filtrates were tested as WAFs. The negative pressure of the filtration unit was reduced as much as possible to avoid losses of volatile components of the test item during filtration.
The stirring period of 3 hours was chosen based on the results of pre-experiments (non-GLP) in which the maximum concentration of the different compounds in the test media was reached after the stirring period of 3 hours.
The test media were prepared just before the start of the test.
- Controls: Test water without test item

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Strain/clone: Daphnia magna Straus clone 5
- Source: A clone of this species (defined by the supplier as clone 5) was originally supplied by the University of Sheffield / UK in 1992. Since that time, the clone has been bred at Harlan Laboratories in reconstituted water of the quality identical to the water quality used in the tests (in respect to pH, main ions, and total hardness) and under temperature and light conditions identical to those of the tests.
- Age: At the start of the test, the organisms used in the test were 6-24 hours old and were not first brood progeny.
- Feeding during test: Daphnids were not fed during the test

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Post exposure observation period:

None

Hardness:

Water Hardness: 250 mg/L as CaCO3

Test temperature:

20 °C

pH:

At the beginning and end of the test period, pH value of the test media was in the range of 7.5 to 7.8

Dissolved oxygen:

At the beginning and end of the test period, the dissolved oxygen concentrations in the test media and control were at least 7.1 mg/L.

Salinity:

No data

Nominal and measured concentrations:

Water accommodated fractions (WAFs) with the loading rates of 4.6, 10, 22, 46 and 100 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 50-mL glass tubes
- Type: closed; the test tubes were made tight with glass stoppers to avoid losses of the volatile substance.
- Material, size, headspace, fill volume: Since the test item was determined to be volatile (according to the results of a pre-experiment, non-GLP), the test was performed in 50-mL glass tubes completely filled with about 50 mL of test medium.
- Aeration: The test water was aerated prior to the start of the study until oxygen saturation was reached. During the test period, the test water was not aerated.
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- A static test without test medium renewal was performed.
- The volume of test solution provided for each daphnia was 10 mL. Thus, the requirement of the test guidelines for the minimum volume of 2 mL test medium per daphnia was fulfilled. The daphnids were randomly distributed to the test vessels at initiation of the test.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Reconstituted test water according to ISO 6341 was used in the study.
- Alkalinity: 0.8 mmol/L
- Ratio of Ca:Mg and Na:K was 4:1 and 10:1, respectively, based on molarity

OTHER TEST CONDITIONS
- Test was performed as far as possible in the dark to avoid a photolytic degradation of the test item.

EFFECT PARAMETERS MEASURED
- Determination of the Immobility: The immobility of the daphnids was determined by visual inspection after 24 and 48 hours of exposure. Those daphnids that were not able to swim within 15 seconds after gentle agitation of the test vessel were considered to be immobilized.
- Water Quality Criteria: At the start and end of the test, the pH values, dissolved oxygen concentrations and water temperature were determined at each test concentration and in the control. The appearance of the test media was visually recorded at the start of the test and after 24 and 48 hours.

RANGE-FINDING STUDY
- The selection of the loading rates was based on the results of a range-finding test and on results of a pre-experiment to determine the solubility of the test item in the test water (non-GLP).

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

48 h

Dose descriptor:

EL50

Effect conc.:

15 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% confidence limits: 10 and 22 mg/L

Remarks:

geometric mean value of the two consecutive test concentrations with 0 and 100% immobility (EL0 and EL100)

Duration:

48 h

Dose descriptor:

other: The highest tested loading rate without observed effect

Effect conc.:

4.6 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- During the first 24 hours of the test, no immobilized test organisms were determined in the control and up to and including the loading rate of 10 mg/L. From the loading rate of 22 mg/L onwards, all daphnids were found to be immobile. Thus, the concentration-effect relationship was very steep. The 24-hour EL50 of the test item was calculated to be 15 mg/L with 95% confidence limits of 10 and 22 mg/L. The 24-hour EL0 was 10 mg/L. The 24-hour EL100 was 22 mg/L.
- Additionally to the immobilization recorded after 24 hours, one daphnid was found to be immobile at the loading rate of 10 mg/L (5% immobility) after 48 hours of exposure. This low immobilization rate was estimated as a significant toxic effect, but had no influence on the calculation of the 48-hour EL50. The 48-hour EL50 was calculated to be 15 mg/L with 95% confidence limits of 10 and 22 mg/L. The 48-hour EL0 and the 48-hour NOEC (highest concentration tested without toxic effects after 48 hours) was 4.6 mg/L, since no significant immobilization was observed up to and including this test concentration. The 48-hour EL100 was 22 mg/L.
- No remarkable observations were made concerning the appearance of the test media. All test media were clear solutions throughout the entire test duration.

Results with reference substance (positive control):

For evaluation of the quality of the daphnia clone and the experimental conditions, potassium dichromate is tested as a positive control twice a year. The result of the latest positive control test in March 2008 (48-hour EC50: 0.73 mg/L, RCC Study No. B83744) indicated that the sensitivity of the test organisms was within the historical range of Harlan Laboratories (48-hour EC50 from 1996 to 2008: 0.53-1.1 mg/L).

Reported statistics and error estimates:

The 24- and 48-hour EL50 could not be calculated by Probit Analysis or Moving Average Interpolation due to the steep concentration-effect relationship. Instead, the 48-hour EL50 was determined as the geometric mean value of the two consecutive test concentrations with 0 and 100% immobility (EL0 and EL100), and the confidence limits for the EL50 as the test concentrations with 0 and 100% immobility.
The NOEC, EL0 and EL100 were determined directly from the raw data.

Table 6.1.3/2: Effect of test item on the Mobility of Daphnia magna

Loading Rate (mg/L)	No. of daphnids tested	Immobilized daphnids after 24 hours		Immobilized daphnids after 48 hours
		No.	%	No.	%
Control	20	0	0	0	0
4.6	20	0	0	0	0
10	20	0	0	1	5
22	20	20	100	20	100
46	20	20	100	20	100
100	20	20	100	20	100

 

At the start of the test, the measured test item concentrations (based on the sum of four peaks) in the test media with the loading rates of 4.6, 10, 22, 46 and 100 mg/L were 0.30, 0.80, 2.1, 3.2 and 4.2 mg/L, at the end of the test 0.17, 0.49, 1.1, 1.3 and 2.2 mg/L, respectively.

Since water accommodated fractions of the test item were tested, all reported biological results were based on the loading rates of the test item.

Validity criteria fulfilled:

yes

Conclusions:

Under the test conditions, the 48-hour EL50 of the test item was calculated to be 15 mg/L (95% confidence limits of 10 and 22 mg/L).

Executive summary:

A study was performed according to OECD Guideline 202 with GLP statement, to assess the 48 h-acute toxicity of the of the test item to Daphnia magna, under static conditions.

 

In order to assess the toxicity of test item containing different components to daphnids, water accommodated fractions (WAFs) with the loading rates of 4.6, 10, 22, 46 and 100 mg/L were tested. Additionally, a control (test water without test item) was tested in parallel.

For preparation of the WAFs, individual dispersions of the test item with the loading rates as mentioned above were prepared. The dispersions were stirred for 3 hours to dissolve a maximum amount of the different compounds of the test item in the dispersion. Then, the dispersions were filtered through membrane filters and the undiluted filtrates were tested as WAFs. The negative pressure of the filtration unit was reduced as much as possible to avoid losses of volatile components of the test item during filtration. The stirring period of 3 hours was chosen based on the results of pre-experiments (non-GLP) in which the maximum concentration of the different compounds in the test media was reached after the stirring period of 3 hours.

 

At the start of the test, the measured test item concentrations (based on the sum of four peaks) in the test media with the loading rates of 4.6, 10, 22, 46 and 100 mg/L were 0.30, 0.80, 2.1, 3.2 and 4.2 mg/L, at the end of the test 0.17, 0.49, 1.1, 1.3 and 2.2 mg/L, respectively. Since water accommodated fractions of the test item were tested, all reported biological results were based on the loading rates of the test item.

 

During the first 24 hours of the test, no immobilized test organisms were determined in the control and up to and including the loading rate of 10 mg/L. From the loading rate of 22 mg/L onwards, all daphnids were found to be immobile. Thus, the concentration-effect relationship was very steep. Additionally to the immobilization recorded after 24 hours, one daphnid was found to be immobile at the loading rate of 10 mg/L (5% immobility) after 48 hours of exposure. This low immobilization rate was estimated as a significant toxic effect, but had no influence on the calculation of the 48-hour EL50. No remarkable observations were made concerning the appearance of the test media. All test media were clear solutions throughout the entire test duration.

 

Under the test conditions, the 48-hour EL50 of the test item was calculated to be 15 mg/L (95% confidence limits of 10 and 22 mg/L).

Reference 3

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

supporting study

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH
[further information is included as attachment to Iuclid section 13]

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
This read-across is based on the hypothesis that source and target substances have similar physico-chemical, environmental fate and (eco)toxicological properties because of their structural and composition similarity.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
See Read-across justification document in Section 13 of IUCLID.

3. ANALOGUE APPROACH JUSTIFICATION
In addition to the structural/composition similarity, physico-chemical and environmental fate properties, both substances have been shown to be toxic to aquatic organisms (all EL50 values are comprised between 1 and 10 mg/L, except the EL50 value on the aquatic invertebrates Daphnia magna for the source substance at 15 mg/L). No experimental study is available for the target substance. The acute toxicity of the target substance to the aquatic invertebrates was determined using QSAR result (iSafeRat® Holistic HA-QSAR v1.7). The QSAR result supports the experimental study performed on the source substance with predicted EL50 value more conservative but in the same order of magnitude. The main components of both substances have the same mode of action (MoA 1, non-polar narcosis) with similar log Kow values. The ecotoxicological properties between the target and source substances are pronounced. Therefore, it’s considered suitable and scientifically justified to read-across the data between the two substances to fill the short-term toxicity to aquatic invertebrates endpoint in the present dossier.

4. DATA MATRIX
See Read-across justification document in Section 13 of IUCLID.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across: supporting information

Key result

Duration:

48 h

Dose descriptor:

EL50

Effect conc.:

15 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% confidence limits: 10 and 22 mg/L

Remarks:

geometric mean value of the two consecutive test concentrations with 0 and 100% immobility (EL0 and EL100)

Duration:

48 h

Dose descriptor:

other: The highest tested loading rate without observed effect

Effect conc.:

4.6 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Validity criteria fulfilled:

not applicable

Conclusions:

According to the experimental study performed on the source substance, the 48-hour EL50 of the target substance can be determined at 15 mg/L (95% confidence limits of 10 and 22 mg/L).

Executive summary:

No experimental study is available for the target substance to assess the short-term toxicity to aquatic invertebrates. However, a study was performed according to OECD Guideline 202 with GLP statement, to assess the 48 h-acute toxicity of a read-across (source) substance to Daphnia magna, under static conditions.

Both substances (the target and the source substances) are UVCB substances with very similar compositions. In the experimental study performed on the source substance, Water Accommodated Fractions (WAFs) with the loading rates of 4.6, 10, 22, 46 and 100 mg/L were tested. Additionally, a control (test water without test item) was tested in parallel.  

At the start of the test, the measured test item concentrations (based on the sum of four peaks) in the test media with the loading rates of 4.6, 10, 22, 46 and 100 mg/L were 0.30, 0.80, 2.1, 3.2 and 4.2 mg/L, at the end of the test 0.17, 0.49, 1.1, 1.3 and 2.2 mg/L, respectively. Since water accommodated fractions of the test item were tested, all reported biological results were based on the loading rates of the test item.

During the first 24 hours of the test, no immobilized test organisms were determined in the control and up to and including the loading rate of 10 mg/L. From the loading rate of 22 mg/L onwards, all daphnids were found to be immobile. Thus, the concentration-effect relationship was very steep. Additionally to the immobilization recorded after 24 hours, one daphnid was found to be immobile at the loading rate of 10 mg/L (5% immobility) after 48 hours of exposure. This low immobilization rate was estimated as a significant toxic effect, but had no influence on the calculation of the 48-hour EL50. No remarkable observations were made concerning the appearance of the test media. All test media were clear solutions throughout the entire test duration.

Under the test conditions, the 48-hour EL50 of the test item was calculated to be 15 mg/L (95% confidence limits of 10 and 22 mg/L).

Description of key information

QSAR, iSafeRat® Holistic HA-QSAR v1.7, calculation method, WAF, key study, validity 1:

48h-EL50 = 5.1 mg/L.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

5.1 mg/L

Additional information

To assess the short-term toxicity of the registered substance to aquatic invertebrates, a set of two data are available.

The first data (KREATiS, 2017), assessed as a key data point, is a QSAR. This QSAR used a calculation method to predict the acute toxicity of the registered substance, an UVCB substance, to aquatic invertebrates Daphnia sp. (48h-EL50 value). This calculation method predicts the endpoint value which would be expected when testing the substance under experimental conditions in a laboratory following OECD Guideline 202 adapted for testing as a mixture using the WAF method. This method has previously been validated in an internal publication (Bicheral and Thomas, 2014; available in the Endpoint Study Record). This algorithm is based on a QSAR model (iSafeRat® Holistic HA-QSAR v1.7) which has been validated to be compliant with the OECD recommendations for QSAR modelling (OECD, 2004). The QSAR is based on validated data for a training set of 58 chemicals derived from 48h test on daphnids, for which the concentrations of the test item had been determined by chemical analyses over the test period. Further to this, the effective loading rate of the WAF is determined by using a series of calculation steps using phase equilibrium thermodynamics and excluding the non-bioavailable fraction. The 48h-EL50 value based on mobility was determined to be 5.1 mg/L.

To support this prediction and reinforce the correct order of magnitude, a second data was provided in the dossier. This data is an experimental study (Harlan, 2009) performed on a source substance, according to OECD Guideline 202 with GLP statement. The 48 h-acute toxicity of this substance to Daphnia magna, under static conditions, was assessed using WAF method. Both substances (the target and the source substances) are UVCB substances with very similar compositions. The main components of both substances have the same mode of action (MoA 1, non-polar narcosis) with similar log Kow values. In this experimental study, the 48-hour EL50 of the test item was calculated to be 15 mg/L (95% confidence limits of 10 and 22 mg/L). This experimental result on the source substance supports the QSAR key result performed on the target substance, with a predicted EL50 value more conservative.

In conclusion, according to the key data, the retained 48h-EL50 value for the registered substance on aquatic invertebrates is 5.1 mg/L.