Hexaphenoxycyclotriphosphazene

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Phenocycycloposphazene, did not exhibit indications of mutagenic potential under the conditions of in vitro Genetic toxicity study.

Link to relevant study records

Reference

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

July 21, 1997

Qualifier:

according to guideline

Guideline:

other: Testing Guidelines for Studies of Chemicals

Version / remarks:

NOtification No. 2018-12 issued by the National Institute of Environmental Research, Republic of Korea on 9 April, 2018

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Specific details on test material used for the study:

Lot No. 180705
Purity: 99.08%
Appearance: White crystal
Storage condition: Room temperature

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Metabolic activation:

with and without

Metabolic activation system:

S9 mix

Test concentrations with justification for top dose:

The high does was selected at 5,000 ug/plate and it was sequentially diluted by the drometric ratio of 4 to produce lower dose levels (1250, 313, 78.1, 19.5 and 4.88 ug/plate).

Vehicle / solvent:

DMSO

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

dissoved in DMSO

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

9-aminoacridine

2-nitrofluorene

sodium azide

benzo(a)pyrene

other: 2-Aminoanthracene (2-AA)

Details on test system and experimental conditions:

The main study was conducted according to the direct plate incorporation method under both absence and presence of metabolic activation system. Three plates per dose were used in the main study and conducted in duplicate. Each plate was labeled with an identification numbers which contained the bacterial strain, dose, the positive ad negative controls and the absence or presence of S9 mix. In the absence of metabolic activation, 100 ul of each test substance, the strain-specific positive control and negative control were placed in the respective tubes. 500 ul of 0.1 mol/L sodium phosphate buffer (pH 7.4) was added and followed by addition of 100 uL of pre-incubated bacterial suspension. Then, 2 ml of warmed top agar for Salmonslla typhimurium was added to TA98, TA100, TA1535 and TA1537 strains and 2mL of warmed top agar for Escherichia coli was added to the WP2uvrA strain. They were mixed thoroughly with a vortex mixer. Finally these mixtures were poured on the minimal glucose agar plates and allowed to solidify at room temperature. In the presence of metabolic activation, S9 mix instead of 500 ul of 0.1 mol/L sodium phosphate buffer (pH7.4) was added, ad the rest of procedure was carried out with the same method as above. In order to confirm microbial contamination, 100 ul of each high dose formulation, 500 uL of 0.1 mol/L sodium phosphate buffer (pH7.4) and 500 uL of S9 mix were placed in the respective tubes. 2 ml of warmed top agar was added and mixed thoroughly with a vortex mixer. Then, the mixed solution was poured. After the op agar was solidified, the plates were inverted and cultured in an incubator at approximately 37 oC for 38 hours.
Following cultivation, the number of revertant colonies was counted by visual and colony counter (postive controls only). Individual plate counts were written for revertant colonies. The average and standard deviations of revertant colonies were calculated. To confirm the presence of growth inhibition and precipitation, the background lawn was observed using a stereoscopic microscope. Cytotoxicity was observed in comparison with the negative control, by a reduction in the number of revertant colonies, diminution of clearing of background lawn compared to the negative control.

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Remarks on result:

no mutagenic potential (based on QSAR/QSPR prediction)

Conclusions:

Based on the results of this study, the test substance, Phenocycycloposphazene, did not exhibit indications of mutagenic potential under the conditions of this study.

Executive summary:

This study was designed to determine the mutagenic potential of Phenoxycycloposphazene using histidine requiring Salmonella typhimurium (TA98, TA100, TA1535 and TA1537) strains and tryptophan requiring Escherichia coli (WP2uvrA) strain.

In order to determine the high dose levels of the main study, the dose range finding study was conducted. The high does was selected at 5,000 ug/plate and it was sequentially diluted by the drometric ratio of 4 to produce lower dose levels (1250, 313, 78.1, 19.5 and 4.88 ug/plate). As a result, the growth inhibition by the test substance was not evident in all strains in presence and absence of metabolic activation. The precipitation by the test substance was evident at 5000 ug/plate in absence and presence of metabolic activation. The precipitation did not interfere with the scoring.

Therefore, the dose levels of the main study was selected as follows. In addition, the positive and negative control groups were set.

Strains	S9 mix	Does levels of the main study (ug/plate)
TA98, TA100, TA1535, TA1537, WP2uvrA	-/+	5000, 2500,1250,625,313

Based on the results of 1st and 2nd main studies, the mean number of revertant colonies in all strains was no differences when compared to that of the negative control group at all dose levels of the test substance in the absence and presence of metabolic activation, without dose-related increase. The growth inhibition by the test substance was not observed under all conditions. The precipitations that did not interfere with the scoring were observed at more than 2500 ug/plate in absence and presence of metabolic activation.

The mean number of revertant colonies for the negative controls is within the range of historic control data. In the positive control group, the number of revertant colonies was markedly increased more than twice when compared to that of the negative control group. Based on the results of this study, the test substance, Phenocycycloposphazene, did not exhibit indications of mutagenic potential under the conditions of this study.

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

Since Phenocycycloposphazene did not exhibit indications of mutagenic potential under the conditions of in vitro Genetic toxicity study, it is not classifed for Genetic toxicity.