Trisodium 8-hydroxypyrene-1,3,6-trisulphonate

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

AMES test

The test chemical was considered  to be  non – mutagenic in the Salmonella – Microsome assay.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.

In vitro gene mutation assay in Mammalian cells

The test chemical was cosidered to be  non – mutagenic in the Chromosomal Aberration assay using a mammalian cell culture

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.

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Justification for type of information:

Data is from publication

Qualifier:

according to guideline

Guideline:

other: As mention below

Principles of method if other than guideline:

To determine the genetic toxicity of the test material by AMES test.

GLP compliance:

not specified

Type of assay:

bacterial gene mutation assay

Target gene:

AMES Assay

Species / strain / cell type:

S. typhimurium, other: No detailed data available

Details on mammalian cell type (if applicable):

Not applicable

Additional strain / cell type characteristics:

not specified

Metabolic activation:

not specified

Metabolic activation system:

no data

Test concentrations with justification for top dose:

no data

Vehicle / solvent:

no data

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

not specified

True negative controls:

not specified

Positive controls:

not specified

Positive control substance:

not specified

Remarks:

no data

Details on test system and experimental conditions:

no data

Evaluation criteria:

An increase in the number of revertants was noted

Statistics:

no data

Species / strain:

S. typhimurium, other: No detailed data available

Metabolic activation:

not specified

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Remarks on result:

other: No mutagenic potential

Table 1 : Genetoxicity of the tracer dyes

Tracer	Salmonella –Microsome assay	Cytogenetic assay – Chromosal aberration
Pyranine	-	-

Conclusions:

The test chemical was considered to be non – mutagenic in the Salmonella – Microsome assay.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.

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. Two test methods that have been sufficiently validated and established for testing of chemicals were used: the salmonella/microsome test (gene mutation) and the analysis of chromosome aberration using a mammalian cell culture (chromosome aberration).The test chemical was considered to be non – mutagenic in the Salmonella – Microsome assay. 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.Test substance is classified as SAFE for use in water tracing.

Reference 2

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Justification for type of information:

Data is from publication.

Qualifier:

according to guideline

Guideline:

other: As mention below

Principles of method if other than guideline:

To determine the genetic toxicity of the test material .

GLP compliance:

not specified

Type of assay:

in vitro mammalian chromosome aberration test

Target gene:

no data

Species / strain / cell type:

mammalian cell line, other: No detailed data available

Details on mammalian cell type (if applicable):

No data

Additional strain / cell type characteristics:

not specified

Metabolic activation:

not specified

Metabolic activation system:

no data

Test concentrations with justification for top dose:

No data

Vehicle / solvent:

no data

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

not specified

True negative controls:

not specified

Positive controls:

not specified

Positive control substance:

not specified

Details on test system and experimental conditions:

no data

Evaluation criteria:

Chromosmal abberration,gaps were noted in cell line.

Statistics:

no data

Species / strain:

mammalian cell line, other:

Metabolic activation:

not specified

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Additional information on results:

No data

Remarks on result:

other: No mutagenic potential was observed

Table 1: Genetic Toxicity of tracer dyes

Tracer	Salmonella Microsome assay	Cytogenetic assay – Chromosal aberration
Pyranine	-	-

Conclusions:

The test chemical was cosidered to be non – mutagenic in the Chromosomal Aberration assay using a mammalian cell culture
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.

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. Two test methods that have been sufficiently validated and established for testing of chemicals were used: the salmonella/microsome test (gene mutation) and the analysis of chromosome aberration using a mammalian cell culture (chromosome aberration).The test chemical was considered to be non – mutagenic in the Salmonella – Microsome assay. 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.Test substance is classified as SAFE for use in water tracing.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Gene mutation in vitro:

Data for the various test chemicals was reviewed to determine the mutagenic nature of Solvent Green 7; Pyranine; trisodium 8-hydroxypyrene-1,3,6-trisulfonate (6358-69-6). The studies are as mentioned below:

AMES test

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. Two test methods that have been sufficiently validated and established for testing of chemicals were used: the salmonella/microsome test (gene mutation) and the analysis of chromosome aberration using a mammalian cell culture (chromosome aberration).The test chemical was considered to be non – mutagenic in the Salmonella – Microsome assay. 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.Test substance is classified as SAFE for use in water tracing.

Supported by other studies.

Ames assay was performed to investigate the potential of the test chemical to induce gene mutations in comparison to negative control according to the plate incorporation test (Trial I) and the pre-incubation test (Trial II) using the Salmonella typhimurium strains TA 1535, TA 1537, TA 98, TA 100 and TA 102. The assay was performed in two independent experiments both with and without liver microsomal activation. Each concentration, including the negative, positive controls was tested in triplicate. Based on the pre-experiment results, the test item was tested with the following concentrations 0.0 (NC), 0.005, 0.016, 0.050, 0.158, 0.501 mg/plate for main study, both in the presence of metabolic activation (+S9) and in the absence of metabolic activation (-S9). No substantial increase in revertant colony numbers in any of the tester strains were observed following treatment with the test chemical at any dose level in both the confirmatory trials, neither in the presence nor in the absence of metabolic activation (S9 mix). In conclusion, it is stated that during the described mutagenicity test and under the experimental conditions reported, the test chemical did not induce gene mutations by base pair changes or frame shifts in the genome of the strains used.  

Gene mutation toxicity study was performed to determine the mutagenic nature of the test chemical. The study was performed using S. typhimurium strains TA98 and TA100 with and without S9 metabolic activation system. The test was performed as per the plate incorporation assay at dose level of 0 (negative control), 50 or 200µg/plate. The chemical was dissolved in water. The result was considered positive when a reproducible, dose-related, at least two-fold increase in the number of revertants over background was observed. Concurrent positive and negative control chemicals were also included in the study. The test chemical did not induce a doubling of revertant colonies over the negative control using S. typhimurium strains TA98 and TA100 in the presence and absence of S9 metabolic activation system and hence the chemical is not likely to classify as a gene mutant in vitro.

In vitro gene mutation assay in Mammalian cells

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. Two test methods that have been sufficiently validated and established for testing of chemicals were used: the salmonella/microsome test (gene mutation) and the analysis of chromosome aberration using a mammalian cell culture (chromosome aberration).The test chemical was considered to be non – mutagenic in the Salmonella – Microsome assay. 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.Test substance is classified as SAFE for use in water tracing.

Supported by other studies.

In vitro mammalian chromosome aberration test was performed to determine the mutagenic nature of the test chemical. The study was performed using CHO-WBL cells in the presence and absence of exogeneous metabolic activation system. The test chemical was dissolved in DMSO and used at dose levels upto 5 mg/mL. In the chromosome aberration assay without activation, cells were exposed to the test chemical for 8 hr. The test chemical was washed off, and the cells were treated with 0.1µg/ml Colcemid for 2-2.5 hr. With metabolic activation, the cells were exposed to the test chemical plus the metabolic activation mixture for 2 hr, washed, incubated for 8 hr, and then treated with Colcemid for 2-2.5 hr. A delayed harvest was used in the aberration assay in most instances when cell cycle delay was observed in the SCE assay. In these tests the cell growth period was extended to about 20 hr. Cells were harvested. Air-dried slides were coded and stained with Giemsa. One hundred to 200 cells from each of the three highest scorable doses were analyzed and the chromosomal aberrations were scored. The test chemical did not induce chromosomal aberrations when tested to toxicity. Precipitate was evident at doses of 250µg/ml and above. Based on the observations made, the test chemical did not induce chromosome aberrations in the CHO-WBL cells in the presence and absence of exogeneous metabolic activation system and hence it is not likely to classify as a gene mutant in vitro.

 

In vitro mammalian chromosome aberration test was performed to determine the mutagenic nature of the test chemical. The study was performed using CHO-WBL cells in the presence and absence of exogeneous metabolic activation system. The test chemical was dissolved in DMSO and used at dose levels upto 5 mg/mL. In the chromosome aberration assay without activation, cells were exposed to the test chemical for 8 hr. The test chemical was washed off, and the cells were treated with 0.1µg/ml Colcemid for 2-2.5 hr. With metabolic activation, the cells were exposed to the test chemical plus the metabolic activation mixture for 2 hr, washed, incubated for 8 hr, and then treated with Colcemid for 2-2.5 hr. A delayed harvest was used in the aberration assay in most instances when cell cycle delay was observed in the SCE assay. In these tests the cell growth period was extended to about 20 hr. Cells were harvested. Air-dried slides were coded and stained with Giemsa. One hundred to 200 cells from each of the three highest scorable doses were analyzed and the chromosomal aberrations were scored. The test chemical did not induce chromosomal aberrations when tested to toxicity. Precipitate was evident at doses of 250µg/ml and above. Based on the observations made, the test chemical did not induce chromosome aberrations in the CHO-WBL cells in the presence and absence of exogeneous metabolic activation system and hence it is not likely to classify as a gene mutant in vitro.

 

Based on the data summarized, Solvent Green 7; Pyranine; trisodium 8-hydroxypyrene-1,3,6-trisulfonate (6358-69-6)did not induce gene mutation .Hence it is not likely to be mutagenic in vitro.

Justification for classification or non-classification

Thus based on the above annotation and CLP criteria the test chemical Solvent Green 7; Pyranine; trisodium 8-hydroxypyrene-1,3,6-trisulfonate (6358-69-6) did not induce gene mutation .Hence it is not likely to be mutagenic in vitro.