Rabitle FP-100

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

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:

key study

Study period:

31st July to 26th September 2003

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: - Study generated according to generally valid and/or internationally accepted testing guidelines - Performed according to GLP - Test parameters based on specific testing guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

Annex V (Ames)

Deviations:

no

Principles of method if other than guideline:

N/A

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Additional strain / cell type characteristics:

not applicable

Species / strain / cell type:

E. coli WP2 uvr A

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with

Metabolic activation system:

Phenobarbital/5,6-benzoflavone induced rat liver S9

Test concentrations with justification for top dose:

Concentration range in the main test (with metabolic activation): 156 ... 5000 µg/plate
Concentration range in the main test (without metabolic activation): 5000 µg/plate

Vehicle / solvent:

Solvent: Dimethylsulphoxide

Details on test system and experimental conditions:

Concentration of the test substance resulting in precipitation: 313 µg/plate

Evaluation criteria:

The test substance was judged positive when the number of revertant colonies increased twice or more than that of the negative control in a dose dependent manner and a reproducibility of the test results was also assured. in other cases, it was judged negative.

Statistics:

None were used

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(> 5000 µg/plate)

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(> 5000 µg/plate)

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

Observations:
No significant increase in the number of revertant colonies
in the test substance treatment groups was observed in any
test strains with or without S9 mix.


The test substance precipitatied at 313 micrograms/plate
without S9 mix and 1250 microgrames/plate with S9 mix.

Bacterial growth inhibition was not observed at any dose
level.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'. Remarks: preliminary test

Conclusions:

Interpretation of results (migrated information):
negative with metabolic activation
negative without metabolic activation

The test substance was judged negative because the number of revertant colonies for the test substance treatment groups in all test strains was less than twice that of the negative control regardless of a presence or absence of S9 mix.
The number of revertant colonies in the positive controls were above two times that of the negative control. The test results showed that the numbers of revertant colonies in the negative control and positive controls were within the range of the historical data at Hita laboratory. It was also confirmed that the test system was free from bacterial contamination, which indicates the test results to be valid.
From the above results, it was concluded that FP-100 had no ability to induce mutations under the present test conditions.

Executive summary:

The ability of FP-100 to induce mutations was investigated by using Salmonella typhimurium strains TA100, TA1535, TA98 and TA1537 and Escherichia coli strain WP2uvrA with a pre-incubation method in the presence of a metabolic activation system (S9 mix).

As a result , the mutagenicity of the test substance was judged negative because the numbers of reventant colonies in the test substance treatment groups were less than two times that of each negative control in all test strains. therfore, it is conlcuded that FP-100 has no ability to induce mutations under the present test conditions.

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:

key study

Study period:

7th October to 27th December 2005

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: - Study generated according to generally valid and/or internationally accepted testing guidelines - Performed according to GLP - Test parameters based on specific testing guideline

Qualifier:

according to guideline

Guideline:

JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals

Deviations:

no

Principles of method if other than guideline:

This study was conducted in accordance with " III Mutagenicity Test: Chromosomal Aberration Test Using Cultured Mamalian cells" prescribed in " Concerning Testing Methods relating to the new chemical substances" (Notification No 1121002 of the Pharmaceutical and Food Safety Bureau, MHLW, No 2 (November 13, 2003) of the Manufacturing industries Bureau, METI & No 031121002 of the Environmental Health Department, MOE (November 21, 2003)

GLP compliance:

yes

Type of assay:

in vitro mammalian chromosome aberration test

Species / strain / cell type:

Chinese hamster lung fibroblasts (V79)

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with

Metabolic activation system:

Phenobarbital/5,6-benzoflavone induced rat liver S9

Test concentrations with justification for top dose:

Concentration range in the main test (with metabolic activation): 158 ... 5000 µg/ml
Concentration range in the main test (without metabolic activation): 1250 ... 5000 µg/ml

Vehicle / solvent:

Acetone

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

acetone

True negative controls:

yes

Positive controls:

yes

Positive control substance:

mitomycin C

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

Acetone

True negative controls:

yes

Positive controls:

yes

Positive control substance:

cyclophosphamide

Details on test system and experimental conditions:

Exposure period (with metabolic activation): 6 hours
Exposure period (without metabolic activation): 6 hours

Fixation time:
18 hours

Evaluation criteria:

The findings were judged to be positive when the frequencies of cells with structural aberrations or numerical aberration cells showed 10% or more with a dose-dependence or the frequencies of cells with chromosomal aberrations showed 5% or more both in the chromosomal aberration test and confirmation test. The other cases were judged to be negative.

Statistics:

No statistical analyses were used.

Species / strain:

Chinese hamster lung fibroblasts (V79)

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

(> 5000 µg/ml)

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

Observations:
Precipitation of the test substance was observed at
78.1 micrograms/mL in the short term tests.

Remarks on result:

other: other:

Remarks:

Migrated from field 'Test system'.

Conclusions:

Interpretation of results (migrated information):
negative with metabolic activation
negative without metabolic activation

In each treatment method, the frequencies of cells with chromosomal aberrations did not fluctuate markedly between two culture dishes, and the frequencies of cells with chromosomal aberrations were below 5% in the negative controls, and the frequencies of cells with structural aberrations excluding gaps were over 20% with positive controls, indicating that the present study was appropriately performed.
In the short-term treatments without and with S9 mix and the continuous treatment, the frequencies of cells with structural aberration cells were below 5% at all observation doses of the test substance. Consequently, the structural aberration and the numerical aberration were judged to be negative.
Based on the above results, it was considered that FP-100 did not possess an abiltiy to induce chromosomal aberrations under the present test conditions.

Executive summary:

The ability of FP-100 to induce chromosomal aberrations was investigated by using Chinese hamster lung fibroblasts (CHL/IU cells). Acetome was used as a solvent.

Based on the results of the cell growth inhibition test, the doses in the chromosomal aberration test were set at 1250, 2500 and 5000 µg/ml in the short term treatment without S9 and in the 24 hour continuous treatment, and 158, 500, 1580 and 5000µg/ml in the short term treatment with S9 mix.

In the chromosomal aberration test, the doses for the observation were selected at 1250, 2500 and 5000 µg/ml in the short term treatments without S9 and in the 24 hour continuous treatment, and at 500, 1580 and 5000µg/ml for the short term treatment with S9 mix. In observation, the frequencies of cells with structural aberrations and numerical cells were scored.

As a result of observation of the specimens, frequencies of cells with structural aberrations or numerical aberration cells showed below 5% at all dosse of the test substance in the short term treatments without and with S9 mix and 24 hours continuous treatment, therefore the structural aberrations and numerical aberations were judged to be negative.

On the other hand the freauencies of cells with structural aberrations and numerical aberration cells in the negative control treated for acetone showed below 5% and the frequencies of cells with structural aberrations in the positive controls treated with mitomycin C and cyclophosphamide showed abvoe 20% indicating the proper performance of the present study.

It is concluded that FP-100 does not possess an ability to induce chromosomal abberations under the present test conditions.

Reference 3

Endpoint:

in vitro gene mutation study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Study period:

11th July to 29th September 2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)

Deviations:

yes

Remarks:

Approximately 5.0 x 106 cells were exposed to designated concentrations of the test item . At a concentration of 50 μg/mL the number of colonies per flask in the toxicity experiment was not evaluable due to a pipetting error.

GLP compliance:

yes

Type of assay:

in vitro mammalian chromosome aberration test

Specific details on test material used for the study:

Name: Rabitle®FP-100
CAS No.: 890525-36-7
Chemical Name: Phenoxyphosphazene oligomer
Batch No.: 150123
Molecular Weight: 693.56-1387.12
Purity: ≥ 99%
Density: 1.322 g/mL
Physical State at RT: powder
Colour: white to slight yellow
Expiry Date: 23 January 2018
Storage Conditions: at room temperature, protected from light

Target gene:

Hprt

Species / strain / cell type:

Chinese hamster lung fibroblasts (V79)

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

Phenobarbital/5,6-benzoflavone induced rat liver S9

Test concentrations with justification for top dose:

The test item was investigated at the following concentrations:
without and with metabolic activation:
1.0, 2.5, 5.0, 10, 25, 50, 100, 250 and 500 μg/mL

Vehicle / solvent:

Tetrahydrofuran (THF)

Untreated negative controls:

yes

Remarks:

Treatment medium

Negative solvent / vehicle controls:

yes

Remarks:

Tetrahydrofuran (THF)

True negative controls:

yes

Positive controls:

yes

Remarks:

Without metabolic activation: EMS; ethylmethanesulfonate; With metabolic activation: DMBA; 7,12-dimethylbenz(a)anthracene

Positive control substance:

ethylmethanesulphonate

Details on test system and experimental conditions:

Exposure period (with metabolic activation): 4 hours
Exposure period (without metabolic activation): 4 hours

Evaluation criteria:

The highest concentration should induce a reduced level of survival of approximately 10-20% relative survival. The lowest concentration should be in the range of the negative control with respect to cell viability and proliferation.
A test chemical is considered to be clearly negative if, in all experimental conditions examined
- none of the test concentrations exhibits a statistically significant increase compared with the concurrent negative control,
- there is no concentration-related increase when evaluated with an appropriate trend-test
- all results are inside the distribution of the historical negative control data.

A test chemical is considered to be clearly positive if, in any of the experimental conditions examined
- at least one of the test concentrations exhibits a statistically significant increase compared with the concurrent negative control, and
- the increase is concentration-related when evaluated with an appropriate trend test, and
- any of the results are outside the distribution of the historical negative control data.
- if there is by chance a low spontaneous mutation rate in the corresponding negative and solvent controls a concentration related increase of the mutations within their range has to be discussed.

Statistics:

Mann-Whitney test.

Key result

Species / strain:

Chinese hamster lung fibroblasts (V79)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

Observations:
Precipitation of the test item was noted at concentrations of 250 μg/mL and higher (with and without metabolic activation).

The test item Rabitle®FP-100 was assessed for its potential to induce gene mutations at the HPRT locus using V79 cells of the Chinese hamster.

The main experiment was carried out without and with metabolic activation. The experiment with metabolic activation was performed by including liver microsomes and NADP for efficient detection of a wide variety of carcinogens requiring metabolic activation.

The selection of the concentrations used in the main experiment was based on data from the pre-experiment according to the OECD guideline 476.

In the main experiment 500 μg/mL was selected as the highest concentration (without and with metabolic activation). The experiment with and without metabolic activation was performed as a 4 h short-term exposure assay. The pH-value detected with the test item was within the physiological range (pH 7.0 ± 0.4).

The test item was investigated at the following concentrations:

without and with metabolic activation:

1.0, 2.5, 5.0, 10, 25, 50, 100, 250 and 500 μg/mL

Precipitation:

Precipitation of the test item was noted at concentrations of 250 μg/mL and higher (with and without metabolic activation).

Toxicity:

No biologically relevant growth inhibition (reduction of relative survival below 70%) was observed after the treatment with the test item in the experiment with and without metabolic activation.

In the experiment without metabolic activation the relative survival was 94% for the highest concentration (500 μg/mL) evaluated . The highest biologically relevant concentration evaluated with metabolic activation was 500 μg/mL with a relative survival of 112% .

Mutagenicity:

In the experiment without and with metabolic activation all validity criteria were met. The mutant values of the negative and solvent controls fall within the historical data range of the test facility and the cloning efficiencies of the negative and solvent controls are > 50%.

The positive controls, DMBA (1.5 μg/mL) and EMS (300 μg/mL) showed statistically significant increases in mutant frequency, thereby demonstrating both the sensitivity and validity of the test systems.

In the experiment without metabolic activation the mutant values of the negative controls, the solvent controls and most mutant values of the test item concentrations found were within the historical control data of the test facility Eurofins Munich (about 9-36 mutants per 106 cells).

The mutant frequencies of the negative controls were 24.57 and 27.09 mutants per 106 cells, of the solvent controls 18.56 and 16.56 mutants per 106.cells, respectively, and in the range of 11.27 to 29.75 mutants per 106 cells with the test item, respectively.

The highest mutant frequency was observed at a concentration of 50 μg/mL (29.75 mutants per 106 cells) with a relative survival of 104%.

A statistical analysis displayed that some of the mutant frequencies were significantly increased over those of the solvent controls, but there was no evidence for a dose-response relationship. Therefore this effect was considered as not biologically relevant.

In the experiment with metabolic activation the mutant values of the negative controls, the solvent controls and most mutant values of the test item concentrations found were within the historical control data of the test facility Eurofins Munich (about 8-37 mutants per 106 cells).

The mutant frequencies of the negative controls were 13.44 and 32.49 mutants per 106 cells, of the solvent controls 29.92 and 20.00 mutants per 106.cells, respectively, and in the range of 16.02 to 34.13 mutants per 106 cells with the test item, respectively.

The highest mutant frequency was observed at a concentration of 2.5 μg/mL (34.13 mutants per 106 cells) with a relative survival of 94%.

The mutant frequencies induced by the test item did not show a biologically relevant increase. None of the observed mutant frequencies were statistically significant increased over those of the solvent controls .

Tables are in the attached report

 

Conclusions:

In conclusion, in the described in vitro cell gene mutagenicity test under the experimental conditions reported, the test item Rabitle®FP-100 is considered to be non-mutagenic in the HPRT locus using V79 cells of the Chinese Hamster.

Executive summary:

The test item Rabitle®FP-100 was assessed for its potential to induce mutations at the HPRT locus using V79 cells of the Chinese Hamster.

The selection of the concentrations was based on data from the pre-experiments. The main experiment with and without metabolic activation was performed as a 4 h short-term exposure assay.

The test item was investigated at the following concentrations:

without and with metabolic activation:

1.0, 2.5, 5.0, 10, 25, 50, 100, 250 and 500 μg/mL

Precipitation of the test item was noted in the main experiment at 250 μg/mL and higher

No growth inhibition was observed in the experiment without and with metabolic activation.

In the experiment no biologically relevant increase of mutants was found after treatment with the test item (without and with metabolic activation). All mutant values are within the historical data base of the test facility.

A statistical analysis displayed that some of the mutant frequencies were significantly increased over those of the negative controls, but there was no evidence for a dose-response relationship.

DMBA and EMS were used as positive controls and showed distinct and biologically relevant effects in mutation frequency.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

• In conclusion, in the described in vitro cell gene mutagenicity test under the experimental conditions reported, the test item Rabitle®FP-100 is considered to be non-mutagenic in the HPRT locus using V79 cells of the Chinese Hamster.
• The ability of FP-100 to induce mutations was investigated by using Salmonella typhimurium strains TA100, TA1535, TA98 and TA1537 and Escherichia coli strain WP2uvrA with a pre-incubation method in the presence of a metabolic activation system (S9 mix). As a result , the mutagenicity of the test substance was judged negative because the numbers of reventant colonies in the test substance treatment groups were less than two times that of each negative control in all test strains. therefore, it is conlcuded that FP-100 has no ability to induce mutations under the present test conditions.
• The ability of FP-100 to induce chromosomal aberrations was investigated by using Chinese hamster lung fibroblasts (CHL/IU cells). Acetome was used as a solvent. Based on the results of the cell growth inhibition test, the doses in the chromosomal aberration test were set at 1250, 2500 and 5000 µg/ml in the short term treatment without S9 and in the 24 hour continuous treatment, and 158, 500, 1580 and 5000µg/ml in the short term treatment with S9 mix. In the chromosomal aberration test, the doses for the observation were selected at 1250, 2500 and 5000 µg/ml in the short term treatments without S9 and in the 24 hour continuous treatment, and at 500, 1580 and 5000µg/ml for the short term treatment with S9 mix. In observation, the frequencies of cells with structural aberrations and numerical cells were scored. As a result of observation of the specimens, frequencies of cells with structural aberrations or numerical aberration cells showed below 5% at all dosse of the test substance in the short term treatments without and with S9 mix and 24 hours continuous treatment, therefore the structural aberrations and numerical aberations were judged to be negative. On the other hand the freauencies of cells with structural aberrations and numerical aberration cells in the negative control treated for acetone showed below 5% and the frequencies of cells with structural aberrations in the positive controls treated with mitomycin C and cyclophosphamide showed abvoe 20% indicating the proper performance of the present study. It is concluded that FP-100 does not possess an ability to induce chromosomal abberations under the present test conditions.