1,3,6,8-Pyrenetetrasulfonic acid, sodium salt

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:

2 December 2018 and 3 December 2018

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

results derived from a valid (Q)SAR model, but not (completely) falling into its applicability domain, with adequate and reliable documentation / justification

Justification for type of information:

1. SOFTWARE
Danish QSAR Database
EPISuite

2. MODEL (incl. version number)
A battery of Leadscope and SciQSAR models
ECOSAR

3. SMILES OR OTHER IDENTIFIERS USED AS INPUT FOR THE MODEL
Smiles: C1=CC2=C3C(=C(C=C2S(=O)(=O)[O-])S(=O)(=O)[O-])C=CC4=C(C=C(C1=C43)S(=O)(=O)[O-])S(=O)(=O)[O-].[Na+].[Na+].[Na+].[Na+]
Substance name: Tetrasodium 1,3,6,8-pyrenetetrasulfonate
CAS: 59572-10-0

4. SCIENTIFIC VALIDITY OF THE (Q)SAR MODEL
KREATiS explanation for Klimisch 2: Final QSAR result is reliable and can reliably replace an experimental result with the OECD Guideline for Testing of Chemicals No. 202, "Daphnia sp., Acute Immobilisation Test".

5. APPLICABILITY DOMAIN
This QSAR model has been designed to be used for regulatory purposes and based on the QSAR results, this report predicts the consensus endpoint value which would be expected when testing the substance under experimental conditions in a laboratory following the OECD Guideline for Testing of Chemicals No. 202, "Daphnia sp., Acute Immobilisation Test".

6. ADEQUACY OF THE RESULT
Based on multiple QSAR models applied, the final Acute Aquatic Toxicity to Daphnids (48h EC50) for Tetrasodium 1,3,6,8-pyrenetetrasulfonate was predicted as 1011.92 mg/L.
The final Acute Aquatic Toxicity to Daphnids (48h EC50) was predicted by applying a consensus method on the reliable results derived for individual models. The final QSAR result can be associated with a Klimisch score: K2

Guideline:

other: REACH Guidance on QSARs R.6

Version / remarks:

OECD (2004) Principles for the validation, for regulatory purposes, of (Quantitative) Structure Activity-Relationship Models, http://www.oecd.org/env/ehs/risk-assessment/oecdquantitativestructureactivityrelationshipsprojectqsars.htm

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

OECD (2004) Principles for the validation, for regulatory purposes, of (Quantitative) Structure Activity-Relationship Models, http://www.oecd.org/env/ehs/risk-assessment/oecdquantitativestructureactivityrelationshipsprojectqsars.htm

Specific details on test material used for the study:

C1=CC2=C3C(=C(C=C2S(=O)(=O)[O-])S(=O)(=O)[O-])C=CC4=C(C=C(C1=C43)S(=O)(=O)[O-])S(=O)(=O)[O-].[Na+].[Na+].[Na+].[Na+]

Key result

Effect conc.:

1 011.92 mg/L

Remarks on result:

other: geometric mean of the predicted values

Effect conc.:

2 mg/L

Remarks on result:

other: Leadscope (via Danish QSAR Database)

Remarks:

Outside of applicability domain

Effect conc.:

498.17 mg/L

Remarks on result:

other: SciQSAR (via Danish QSAR Database)

Remarks:

Outside of applicabilit domain

Effect conc.:

1 040 000 mg/L

Remarks on result:

other: ECOSAR (via EPISuite)

Remarks:

neutral organics-acid model

Conclusions:

The final Acute Aquatic Toxicity to Daphnids (48h EC50) predicted for Tetrasodium 1,3,6,8-pyrenetetrasulfonate assigned by the study investigator: 1011.92 mg/L.
Klimisch score assigned by the study investigator for the final prediction: K2

Executive summary:

Introduction. Multiple Quantitative Structure Activity Relationship (QSAR) models were used to predict the Acute Aquatic Toxicity to Daphnids (48h EC50) of the test item Tetrasodium 1,3,6,8-pyrenetetrasulfonate. These QSAR models have been designed to be used for regulatory purposes and based on the QSAR results, this report predicts the consensus endpoint value which would be expected when testing the substance under experimental conditions in a laboratory following the OECD Guideline for Testing of Chemicals No. 202, "Daphnia sp., Acute Immobilisation Test".

 

Methods. The purpose of the in silico study was to predict the Acute Aquatic Toxicity to Daphnids (48h EC50) of the test item Tetrasodium 1,3,6,8-pyrenetetrasulfonate. This prediction was performed using the following QSAR models.

 

·       A battery of Leadscope and SciQSAR models incorporated into Danish QSAR Database

 

·       ECOSAR (EPISuite)

 

 

Results.

Based on multiple QSAR models applied, the final Acute Aquatic Toxicity to Daphnids (48h EC50) for Tetrasodium 1,3,6,8-pyrenetetrasulfonate was predicted as 1011.92 mg/L.

The final Acute Aquatic Toxicity to Daphnids (48h EC50) was predicted by applying a consensus method on the reliable results derived for individual models. The final QSAR result can be associated with a Klimisch score: K2

Reference 2

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

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

Remarks:

Using QSAR

Adequacy of study:

supporting study

Study period:

2 December 2018 and 3 December 2018

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

test procedure in accordance with national standard methods with acceptable restrictions

Justification for type of information:

QSAR information

1. SOFTWARE
Danish QSAR Database
EPISuite

2. MODEL (incl. version number)
A battery of Leadscope and SciQSAR models
ECOSAR

3. SMILES OR OTHER IDENTIFIERS USED AS INPUT FOR THE MODEL
Smiles: [Na+].[Na+].[Na+].OC1=C2C=CC3=C(C=C(C4=CC=C(C(=C1)S([O-])(=O)=O)C2=C34)S([O-])(=O)=O)S([O-])(=O)=O
Substance name: trisodium 8-hydroxypyrene-1,3,6-trisulfonate
CAS: 6358-69-6

4. SCIENTIFIC VALIDITY OF THE (Q)SAR MODEL
KREATiS explanation for Klimisch 2: Final QSAR result is reliable and can reliably replace an experimental result with the OECD Guideline for Testing of Chemicals No. 202, "Daphnia sp., Acute Immobilisation Test".

5. APPLICABILITY DOMAIN
This QSAR model has been designed to be used for regulatory purposes and based on the QSAR results, this report predicts the consensus endpoint value which would be expected when testing the substance under experimental conditions in a laboratory following the OECD Guideline for Testing of Chemicals No. 202, "Daphnia sp., Acute Immobilisation Test".

6. ADEQUACY OF THE RESULT
Based on multiple QSAR models applied, the final Acute Aquatic Toxicity to Daphnids (48h EC50) for Tetrasodium 1,3,6,8-pyrenetetrasulfonate was predicted as 466.73 mg/L.
The final Acute Aquatic Toxicity to Daphnids (48h EC50) was predicted by applying a consensus method on the reliable results derived for individual models. The final QSAR result can be associated with a Klimisch score: K2

-------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------

Read-across information

REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
STRUCTURAL SIMILARITY
Both the substances share the same carbon skeleton, which is pyrene. The query substance holds four sulfonates groups while the proposed analogue has one hydroxy group replacing a sulfonate. Both sulfonate and hydroxy groups are hydrophilic in nature. However, the hydroxy group is only ionised at high pHs (pKa of phenol is around 10) while the sulfonate group would remain ionised over the whole range of aqueous pHs. Therefore, Read-Across substance is expected to be slightly less hydrophilic and more volatile than the query substance, which is confirmed by the log KOW and vapour pressure studies.
Both the substances are expected to be stable in pure form or in water, and they are expected to be not volatile (high boiling point, low vapour pressure), highly hydrophilic (low log KOW, high water solubility) due to the presence of four strongly hydrophilic groups.
 
MECHANISMS OF ACTION PREDICTION
The mechanisms of action of both substances are predicted as follows:

Substance MechoA MechoA detail
Query 1.1 non-polar narcosis for all species
Read-Across 1.2 polar narcosis for all species

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
See Test material sections of the source and target records for details.

3. ANALOGUE APPROACH JUSTIFICATION
DISCUSSION AND CONCLUSION
Therefore, both the structures are expected to have very similar (eco)toxicological profile. Moreover, Read-Across substance being less hydrophilic than the query substance and having a more toxic MechoA, the prediction of (eco)toxicological endpoints will be a worst-case scenario, because the less hydrophilic a substance is, the more toxic it is.
Consequently, trisodium 8-hydroxypyrene-1,3,6-trisulfonate is judged to be a good Read-Across for tetrasodium 1,3,6,8-pyrenetetrasulfonate.

Guideline:

other: REACH Guidance on QSARs R.6

Version / remarks:

OECD (2004) Principles for the validation, for regulatory purposes, of (Quantitative) Structure Activity-Relationship Models, http://www.oecd.org/env/ehs/risk-assessment/oecdquantitativestructureactivityrelationshipsprojectqsars.htm

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

OECD (2004) Principles for the validation, for regulatory purposes, of (Quantitative) Structure Activity-Relationship Models, http://www.oecd.org/env/ehs/risk-assessment/oecdquantitativestructureactivityrelationshipsprojectqsars.htm

Specific details on test material used for the study:

[Na+].[Na+].[Na+].OC1=C2C=CC3=C(C=C(C4=CC=C(C(=C1)S([O-])(=O)=O)C2=C34)S([O-])(=O)=O)S([O-])(=O)=O

Key result

Effect conc.:

466.73 mg/L

Remarks on result:

other: geometric mean of the predicted values

Effect conc.:

500.3 mg/L

Remarks on result:

other: Experimental data from the ECHA dossier: https://echa.europa.eu/registration-dossier/-/registered-dossier/17360/6/2/4/?documentUUID=8939d54a-8147-443c-b2b8-d2b544e7ba11

Effect conc.:

8.69 mg/L

Remarks on result:

other: Leadscope (via Danish QSAR Database)

Remarks:

Outside of applicability domain

Effect conc.:

1 640.5 mg/L

Remarks on result:

other: SciQSAR (via Danish QSAR Database)

Remarks:

Outside of applicabilit domain

Effect conc.:

7 131.724 mg/L

Remarks on result:

other: ECOSAR (via EPISuite)

Remarks:

neutral organics-acid model

Conclusions:

The final Acute Aquatic Toxicity to Daphnids (48h EC50) predicted for trisodium 8-hydroxypyrene-1,3,6-trisulfonate assigned by the study investigator: 466.73 mg/L.
Klimisch score assigned by the study investigator for the final prediction: K2

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

Study period:

March 2011

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

secondary literature

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
STRUCTURAL SIMILARITY
Both the substances share the same carbon skeleton, which is pyrene. The query substance holds four sulfonates groups while the proposed analogue has one hydroxy group replacing a sulfonate. Both sulfonate and hydroxy groups are hydrophilic in nature. However, the hydroxy group is only ionised at high pHs (pKa of phenol is around 10) while the sulfonate group would remain ionised over the whole range of aqueous pHs. Therefore, Read-Across substance is expected to be slightly less hydrophilic and more volatile than the query substance, which is confirmed by the log KOW and vapour pressure studies.
Both the substances are expected to be stable in pure form or in water, and they are expected to be not volatile (high boiling point, low vapour pressure), highly hydrophilic (low log KOW, high water solubility) due to the presence of four strongly hydrophilic groups.
 
MECHANISMS OF ACTION PREDICTION
The mechanisms of action of both substances are predicted as follows:

Substance MechoA MechoA detail
Query 1.1 non-polar narcosis for all species
Read-Across 1.2 polar narcosis for all species

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
See Test material sections of the source and target records for details.

3. ANALOGUE APPROACH JUSTIFICATION
DISCUSSION AND CONCLUSION
Therefore, both the structures are expected to have very similar (eco)toxicological profile. Moreover, Read-Across substance being less hydrophilic than the query substance and having a more toxic MechoA, the prediction of (eco)toxicological endpoints will be a worst-case scenario, because the less hydrophilic a substance is, the more toxic it is.
Consequently, trisodium 8-hydroxypyrene-1,3,6-trisulfonate is judged to be a good Read-Across for tetrasodium 1,3,6,8-pyrenetetrasulfonate.

Guideline:

other: Thain JE, “Biological effects of contaminants: Oyster (Crassostrea gigas) embryo bioassay, Proceedings, Techniques in Marine Environmental Sciences”, International Council for the Exploration of the Sea, 11, pp. 1-12, 1991.

Version / remarks:

http://www.ices.dk/pubs/times/times11/TIMES11.pdf

Deviations:

not specified

GLP compliance:

not specified

Specific details on test material used for the study:

- SMILES: [Na+].[Na+].[Na+].OC1=C2C=CC3=C(C=C(C4=CC=C(C(=C1)S([O-])(=O)=O)C2=C34)S([O-])(=O)=O)S([O-])(=O)=O
- Source of test material: Sigma-Aldrich Co
- purity: greater than 97%
- Colour: Bright yellow Green powder / Bright Green
- Solubility and stability of the test substance in the solvent/vehicle: Clear/good
- Use: Stain or Dye other names: HPTS, Pyranine, Solvent Green 7, 8-Hydroxypyrene-1,3,6-trisulfonic acid trisodium salt

Details on test solutions:

Preparation of tracer dyes
Stock solutions of test dyes were prepared using filtered aerated seawater (0.2 μm). Serial dilutions were then prepared from the main stock to produce the test concentration range.
Initial tests, or range finders, were carried out on both Tisbe battagliai and Crassosterea gigas. The range finder tests covered a concentration range of 0.001 to 1000 mg/L, this was increased to 4000mg/L for tracer dye 8-Hydroxypyrene-1,3,6-trisulfonic acid trisodium salt (Pyranine). The observations allowed for a narrower range to be achieved for more focussed definitive studies.

Test organisms (species):

other: Crassostrea gigas

Water media type:

saltwater

Total exposure duration:

24 h

Test temperature:

24°C ± 2°C

pH:

7.8-8.2

Dissolved oxygen:

equal to or > 80%

Salinity:

32 ± 2 %

Details on test conditions:

The oyster embryo bioassay (OEB) followed the method outlined in Thain JE, “Biological effects of contaminants: Oyster (Crassostrea gigas) embryo bioassay, Proceedings, Techniques in Marine Environmental Sciences”, International Council for the Exploration of the Sea, 11, pp. 1-12, 1991, with no further modifications. These bioassays were carried out in 12 well polystyrene plates, each well contained 4mls of tracer test solution and approximately 50 oyster embryo per ml. The oyster embryos were observed using a high powered binocular microscope and normal and abnormal embryos were scored.

Reference substance (positive control):

yes

Remarks:

Zinc sulphate

Key result

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

485.37 mg/L

Basis for effect:

other: embryo development

Remarks on result:

other: Data generated using range finder results and not true definitives

Details on results:

EC50 values for the oyster embryo bioassay (OEB) were obtained for 10 of the test tracer dyes. The results for Eosin Y, Fluorescein, Rhodamine WT and Sulforhodamine B were determined from definitive tests (with a more defined range etc), whereas the results for other 6 tracers were determined only from the sighting shot test with a broader range of test concentrations. Full definitive tests were not finalised for all the dyes in the OEB tests, due to the unavailability of high quality oysters during all of the testing phase, but the sighting shot results allowed their toxicity to be ranked appropriately for the purposes of this study.
Overall the OEB was a more sensitive test, producing EC50 values that were routinely substantially lower than that of T.battagliai, apart from tracer Rhodamine B and Lissamine. Results revealed the most toxic dye in both tests, with LC50 values below 1mg/l, was Rhodamine 6G. The least toxic was tracer 8-Hydroxypyrene-1,3,6-trisulfonic acid (Pyranine).
Behrens et al (2001) indicated that certain tracer dyes are toxicologically safe and could be used in the marine environment, this included Pryanine.

Validity criteria fulfilled:

yes

Conclusions:

The least toxic was tracer dye 8-Hydroxypyrene-1,3,6-trisulfonic acid (Pyranine): EC50 485.37 mg/L

Executive summary:

The aim of this investigation was to develop a toxicity-based ranked list of the most commonly used fluorescent dyes, with possible applications as marine tracers, as underpinning evidence/support to the marine consents approval process under the Food and

Environment Protection Act (FEPA). Twelve tracer dyes were selected and prioritised using literature and other available data sources. Toxicity tests on the dyes were carried out using two standard bioassays using the copepod acute toxicity test (Tisbe battagliai 48 hr LC50)

and the oyster embryo development bioassay (Crassostrea gigas 24 hr EC50).

Median lethal toxicity (EC50) values were generated for twelve tracer dyes for T.battagliai and 10 tracer dyes for C.gigas. Results showed a broad range of EC50 values across the tracer dyes for both species. EC50 values ranged from 0.0008 to 485 mg/L for oyster and from 0.1 to 2700 mg/L for Tisbe.

The results allowed the ranking of tracer dyes in terms of their toxicity to the two standard test species. The three most toxic tracer dyes were, Rhodamine 6G, Rhodamine B and Lissamine and the least toxic were 8-Hydroxypyrene-1,3,6-trisulfonic acid (Pyranine), Rhodamine WT, Sulforhodamine G, Erioglaucine, Fluorescein.

This ranked list, and supplementary data from the literature, will enable regulatory assessors to determine which tracer dyes pose the highest risk to the aquatic environment more comprehensively and, consequently, strengthen the regulatory advice they provide. Furthermore the data will inform the agreement of licence exemption criteria for tracer dye applications in the marine environment.

Reference 4

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

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

Adequacy of study:

supporting study

Study period:

March 2011

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

secondary literature

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
STRUCTURAL SIMILARITY
Both the substances share the same carbon skeleton, which is pyrene. The query substance holds four sulfonates groups while the proposed analogue has one hydroxy group replacing a sulfonate. Both sulfonate and hydroxy groups are hydrophilic in nature. However, the hydroxy group is only ionised at high pHs (pKa of phenol is around 10) while the sulfonate group would remain ionised over the whole range of aqueous pHs. Therefore, Read-Across substance is expected to be slightly less hydrophilic and more volatile than the query substance, which is confirmed by the log KOW and vapour pressure studies.
Both the substances are expected to be stable in pure form or in water, and they are expected to be not volatile (high boiling point, low vapour pressure), highly hydrophilic (low log KOW, high water solubility) due to the presence of four strongly hydrophilic groups.
 
MECHANISMS OF ACTION PREDICTION
The mechanisms of action of both substances are predicted as follows:

Substance MechoA MechoA detail
Query 1.1 non-polar narcosis for all species
Read-Across 1.2 polar narcosis for all species

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
See Test material sections of the source and target records for details.

3. ANALOGUE APPROACH JUSTIFICATION
DISCUSSION AND CONCLUSION
Therefore, both the structures are expected to have very similar (eco)toxicological profile. Moreover, Read-Across substance being less hydrophilic than the query substance and having a more toxic MechoA, the prediction of (eco)toxicological endpoints will be a worst-case scenario, because the less hydrophilic a substance is, the more toxic it is.
Consequently, trisodium 8-hydroxypyrene-1,3,6-trisulfonate is judged to be a good Read-Across for tetrasodium 1,3,6,8-pyrenetetrasulfonate.

Guideline:

other: ISO, 1999. ISO 14669:1999(E) Water quality -- Determination of acute lethal toxicity to marine copepods (Copepoda, Crustacea)

Deviations:

not specified

GLP compliance:

not specified

Specific details on test material used for the study:

- SMILES: [Na+].[Na+].[Na+].OC1=C2C=CC3=C(C=C(C4=CC=C(C(=C1)S([O-])(=O)=O)C2=C34)S([O-])(=O)=O)S([O-])(=O)=O
- Source of test material: Sigma-Aldrich Co
- purity: greater than 97%
- Colour: Bright yellow Green powder / Bright Green
- Solubility and stability of the test substance in the solvent/vehicle: Clear/good
- Use: Stain or Dye other names: HPTS, Pyranine, Solvent Green 7, 8-Hydroxypyrene-1,3,6-trisulfonic acid trisodium salt

Details on test solutions:

Preparation of tracer dyes
Stock solutions of test dyes were prepared using filtered aerated seawater (0.2 μm). Serial dilutions were then prepared from the main stock to produce the test concentration range.
Initial tests, or range finders, were carried out on both Tisbe battagliai and Crassosterea gigas. The range finder tests covered a concentration range of 0.001 to 1000 mg/L, this was increased to 4000mg/L for tracer dye 8-Hydroxypyrene-1,3,6-trisulfonic acid trisodium salt (Pyranine). The observations allowed for a narrower range to be achieved for more focussed definitive studies.

Test organisms (species):

other aquatic crustacea: Tisbe battagliai

Water media type:

saltwater

Total exposure duration:

48 h

Test temperature:

21°C ± 3°C

pH:

7.8-8.2

Dissolved oxygen:

equal to or > 80%

Salinity:

32 ± 2 %

Details on test conditions:

The T.battagliai bioassay followed the guidelines in ISO (1999) ISO 14669 Water Quality - Determination of acute lethal toxicity to marine copepods (Copepoda, Crustacea), with no further modifications. These bioassays were carried out in 12 well polystyrene plates, each well contained 5mls of test solution and 5 T.battagliai copepodites (approximately 4-6 days old). There were 4 replicates per concentration and the test was carried
out over a period of 48 hours. The Tisbe were observed using a binocular microscope and mortality recorded.

Reference substance (positive control):

yes

Remarks:

Zinc sulphate

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

2 703.1 mg/L

Basis for effect:

mortality

Details on results:

All T.battagliai bioassays were successful in the determination of a definitive EC50 value. The 12 selected tracer dyes exhibited a broad range of EC50 values ranging from 0.1mg/l for Rhodamine 6G, being the most toxic, to 2703.1mg/l for 8-Hydroxypyrene-1,3,6-trisulfonic acid trisodium salt, being the least toxic.
The results for 8-Hydroxypyrene-1,3,6-trisulfonic acid trisodium salt were determined only from the sighting shot test with a broader range of test concentrations.
The least toxic was tracer 8-Hydroxypyrene-1,3,6-trisulfonic acid (Pyranine).
The 5 dyes ranked the least toxic of the 12 were Fluorescein, Erioglaucine, Sulforhodamine G, Rhodamine WT and 8-Hydroxypyrene-1,3,6-trisulfonic acid (Pryanine).
Behrens et al (2001) indicated that certain tracer dyes are toxicologically safe and could be used in the marine environment, this included Pryanine.

Validity criteria fulfilled:

yes

Conclusions:

The least toxic was tracer dye 8-Hydroxypyrene-1,3,6-trisulfonic acid (Pyranine): EC50 2703.1 mg/L

Executive summary:

The aim of this investigation was to develop a toxicity-based ranked list of the most commonly used fluorescent dyes, with possible applications as marine tracers, as underpinning evidence/support to the marine consents approval process under the Food and

Environment Protection Act (FEPA). Twelve tracer dyes were selected and prioritised using literature and other available data sources. Toxicity tests on the dyes were carried out using two standard bioassays using the copepod acute toxicity test (Tisbe battagliai 48 hr LC50)

and the oyster embryo development bioassay (Crassostrea gigas 24 hr EC50).

Median lethal toxicity (EC50) values were generated for twelve tracer dyes for T.battagliai and 10 tracer dyes for C.gigas. Results showed a broad range of EC50 values across the tracer dyes for both species. EC50 values ranged from 0.0008 to 485 mg/L for oyster and from 0.1 to 2700 mg/L for Tisbe.

The results allowed the ranking of tracer dyes in terms of their toxicity to the two standard test species. The three most toxic tracer dyes were, Rhodamine 6G, Rhodamine B and Lissamine and the least toxic were 8-Hydroxypyrene-1,3,6-trisulfonic acid (Pyranine), Rhodamine WT, Sulforhodamine G, Erioglaucine, Fluorescein.

This ranked list, and supplementary data from the literature, will enable regulatory assessors to determine which tracer dyes pose the highest risk to the aquatic environment more comprehensively and, consequently, strengthen the regulatory advice they provide. Furthermore the data will inform the agreement of licence exemption criteria for tracer dye applications in the marine environment.

Reference 5

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

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

Adequacy of study:

supporting study

Study period:

17 March 2001

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

secondary literature

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
STRUCTURAL SIMILARITY
Both the substances share the same carbon skeleton, which is pyrene. The query substance holds four sulfonates groups while the proposed analogue has one hydroxy group replacing a sulfonate. Both sulfonate and hydroxy groups are hydrophilic in nature. However, the hydroxy group is only ionised at high pHs (pKa of phenol is around 10) while the sulfonate group would remain ionised over the whole range of aqueous pHs. Therefore, Read-Across substance is expected to be slightly less hydrophilic and more volatile than the query substance, which is confirmed by the log KOW and vapour pressure studies.
Both the substances are expected to be stable in pure form or in water, and they are expected to be not volatile (high boiling point, low vapour pressure), highly hydrophilic (low log KOW, high water solubility) due to the presence of four strongly hydrophilic groups.
 
MECHANISMS OF ACTION PREDICTION
The mechanisms of action of both substances are predicted as follows:

Substance MechoA MechoA detail
Query 1.1 non-polar narcosis for all species
Read-Across 1.2 polar narcosis for all species

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
See Test material sections of the source and target records for details.

3. ANALOGUE APPROACH JUSTIFICATION
DISCUSSION AND CONCLUSION
Therefore, both the structures are expected to have very similar (eco)toxicological profile. Moreover, Read-Across substance being less hydrophilic than the query substance and having a more toxic MechoA, the prediction of (eco)toxicological endpoints will be a worst-case scenario, because the less hydrophilic a substance is, the more toxic it is.
Consequently, trisodium 8-hydroxypyrene-1,3,6-trisulfonate is judged to be a good Read-Across for tetrasodium 1,3,6,8-pyrenetetrasulfonate.

Qualifier:

equivalent or similar to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

DIN 38412, Part 11 (Deutsches Institut für Normung e.V. 1982)

Specific details on test material used for the study:

[Na+].[Na+].[Na+].OC1=C2C=CC3=C(C=C(C4=CC=C(C(=C1)S([O-])(=O)=O)C2=C34)S([O-])(=O)=O)S([O-])(=O)=O

Key result

Duration:

48 h

Dose descriptor:

EC0

Effect conc.:

>= 10 mg/L

Basis for effect:

mortality

Details on results:

Fluorescent dyes that showed no effect upon either the genotoxicity or the ecotoxicity tests were classified by the Working Group as safe for use in water tracing.

Conclusions:

Pyranine showed no effect in the ecotoxicity tests and was classified by the Working Group as safe for use in water tracing.

Executive summary:

Uncertainties regarding possible negative effects on the environment or on human health of authorizing tracing experiments in groundwater and surface waters led to the establishment of a Working Group at the German Federal Environmental Agency (Umweltbundesamt

– UBA) for conducting a toxicological and ecotoxicological assessment.

A total of 17 water tracers was assessed by the Working Group on the basis of the results of toxicological tests, the available literature, and the group’s expert knowledge. In the future, tracers that pose a risk to the environment or to human health should no longer be used. Nevertheless, there are a number of tracers that could be used in hydrogeological and hydrological investigations for water-pollution-control purposes with no adverse environmental impact.

Description of key information

Multiple Quantitative Structure Activity Relationship (QSAR) models were used to predict the Acute Aquatic Toxicity to Daphnids (48h EC50) of the test item Tetrasodium 1,3,6,8-pyrenetetrasulfonate. The final Acute Aquatic Toxicity to Daphnids (48h EC50) predicted for Tetrasodium 1,3,6,8-pyrenetetrasulfonate assigned by the study investigator: 1011.92 mg/L. Klimisch score assigned by the study investigator for the final prediction: K2

The same method was used to predict the Acute Aquatic Toxicity to Daphnids (48h EC50) of the analogue test item trisodium 8-hydroxypyrene-1,3,6-trisulfonate. The final Acute Aquatic Toxicity to Daphnids (48 EC50) predicted for trisodium 8-hydroxypyrene-1,3,6-trisulfonate assigned by the study investigator: 466.73 mg/L. Klimisch score assigned by the study investigtor for the final prediction: K2

Test results published on the ECHA website provided a conclusion of 500.3 mg/L for Acute Aquatic Toxicity to Daphnids (48h EC50) for the analogue test itemtrisodium 8-hydroxypyrene-1,3,6-trisulfonate

A study was carried out to assess the toxicity of the analogue test item trisodium 8-hydroxypyrene-1,3,6-trisulfonate on Tisbe battagliai (48 hr LC50) and Crassostrea gigas (24 hr EC50) with the following results:

Tisbe battagliai (48 hr LC50): 2703.1 mg/L

Crassostrea gigas (24 hr EC50): 485.37 mg/L

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

1 011.92 mg/L

Additional information

Both structures for the substances Tetrasodium 1,3,6,8-pyrenetetrasulfonate and 8-hydroxypyrene-1,3,6-trisulfonate are expected to have very similar (eco)toxicological profiles. Moreover, the Read-Across substance being less hydrophilic than the query substance and having a more toxic MechoA, the prediction of (eco)toxicological endpoints will be a worst-case scenario, because the less hydrophilic a substance is, the more toxic it is.

Consequently, trisodium 8-hydroxypyrene-1,3,6-trisulfonate is judged to be a good Read-Across for tetrasodium 1,3,6,8-pyrenetetrasulfonate.

Although not completely within the applicability domain, comparison with the QSAR results for the analogue substance, together with the available data on the analogue substance indicate that testing using 8-hydroxypyrene-1,3,6-trisulfonate would not be expected to show evidence of toxicity.

Conclusion: Using Multiple Quantitative Structure Activity Relationship (QSAR) models to predict the Acute Aquatic Toxicity to Daphnids (48h EC50) of the test item Tetrasodium 1,3,6,8-pyrenetetrasulfonate, the final Acute Aquatic Toxicity to Daphnids (48h EC50) predicted for Tetrasodium 1,3,6,8-pyrenetetrasulfonate was 1011.92 mg/L. Although not completely within the compatability domain and taking into consideration the data available on the analogue substance, it can be concluded that under test conditions a 48h EC50 of > 100 mg/L can be expected. Based on this, the test material Tetrasodium 1,3,6,8-pyrenetetrasulfonate is not considered to be toxic to Daphnids at < 100 mg/L.