2-[(2-hydroxyphenyl)methyl]phenol; 2-[(4-hydroxyphenyl)methyl]phenol; 4-[(4-hydroxyphenyl)methyl]phenol

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

7 October 2008

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

2009

Report date:

2008

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

2-[(2-hydroxyphenyl)methyl]phenol; 2-[(4-hydroxyphenyl)methyl]phenol; 4-[(4-hydroxyphenyl)methyl]phenol

EC Number:

908-912-9

Cas Number:

1333-16-0

Molecular formula:

C39H36O6

IUPAC Name:

2-[(2-hydroxyphenyl)methyl]phenol; 2-[(4-hydroxyphenyl)methyl]phenol; 4-[(4-hydroxyphenyl)methyl]phenol

Test material form:

solid

Method

Target gene:

Histiene locus

Species / strain

Species / strain 1

Species / strain / cell type:

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

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A pKM 101

Metabolic activation:

with and without

Metabolic activation system:

S9 mix

Test concentrations with justification for top dose:

0.5, 5, 50, 250, 500 µg.
All compounds were tested at the limit of solubility.

Vehicle / solvent:

DMSO
BPF was dissolved in DMSO and then added to the culture medium such as the final concentration of DMSO did not exceed 0.1% (v/v). All compounds were
tested at the limit of solubility.

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Remarks:

1 µg/plate fo TA98

Positive control substance:

2-nitrofluorene

Remarks:

Without metabolic activation

Controls 2

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Remarks:

2 µg/plate for TA100 and TA1535

Positive control substance:

sodium azide

Remarks:

Without metabolic activation

Controls 3

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Remarks:

1 µg/plate for TA1537

Positive control substance:

other: Acridine Mutagen ICR 191

Remarks:

Without metabolic activation

Controls 4

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Remarks:

0.5 µg/plate for WP2 uvrA

Positive control substance:

4-nitroquinoline-N-oxide

Remarks:

without metabolic activation

Controls 5

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Remarks:

2.5 µg/plate for TA98

Positive control substance:

benzo(a)pyrene

Remarks:

With metabolic activation

Controls 6

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Remarks:

2.5 µg/plate fpt TA100, TA1535, TA1537, WP2 uvrA

Positive control substance:

other: 2-aminoanthracene

Remarks:

with metabolic activation

Details on test system and experimental conditions:

The Ames test was carried out using the plate incorporation method (with preincubation) with or without metabolic activation, with four histidine-dependent auxotrophic mutants of Salmonella typhimurium strains, TA 98, TA 100, TA 1535, TA 1537, essentially as described by Maron and Ames (1983). A fifth strain was used, a tryptophan-dependent auxotrophic mutant Escherichia coli WP2 uvrA pKM101. The test strains were cultured in the liquid broth medium for 10 h at 37 °C under agitation. After incubation, 0.5 ml of 0.1M sodium phosphate buffer (pH 7.4) (absence of metabolic activation) or 0.5 ml of S9 mix (presence of metabolic activation), 0.1ml of bacterial culture and 50 ml of BPF solutions (0.01mg/ml up to 10mg/ml) were added to a test tube and preincubated 1 h at 37 °C. Two millilitre of semi-liquid superficial agar was added to the mixture and poured onto a minimal glucose agar plate. For experiments with S. typhimurium, the top agar was supplemented with 10 ml of 0.5mM histidine/biotin solution per 100ml agar, and mutations to histidine independence were scored on minimal glucose agar plates. For experiments with E. Coli strain, mutations to tryptophan independence were scored on minimal glucose agar plates supplemented with 10 ml of 0.5mM tryptophan per 100ml agar. The plates were incubated 48 h at 37 °C, and then the number of revertant colonies was counted. All experiments were carried out in triplicate using five concentrations of BPF. Mutagenic activities were expressed as induction factors, i.e. as multiples of the background levels.

Evaluation criteria:

According to the historical values of the laboratory, a compound tested with the Ames test was considered mutagenic if the number of His+ revertant
colonies was at least twice the value of the corresponding solvent control (induction factor > 2). A dose-effect relationship is an additional indication for the mutagenic potency of a molecule. A possible mutagenic potential is assumed if the quotient ranges between 1.7 and 1.9 in combination with an observed dose-effect
relationship. No mutagenic potential is assumed if all quotients range between 1.0 (or lower) and 1.6. The latter conclusions can be strengthend by the lack of
observation of a dose-effect relationship.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at highest concentration of 500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at highest concentration of 500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at highest concentration of 500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at highest concentration of 500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 5

Species / strain:

E. coli WP2 uvr A pKM 101

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at highest concentration of 500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

BPF was cytotoxic for the bacteria at the highest concentration tested (500 µg/plate), especially without exogenous activation system. In this study, none of the results of the Ames test (+S9 or −S9) exceeded the critical value of 2.0 and all quotients ranged below 1.6, therefore the Ames test did not show any genotoxic potential of BPF, compared to the respective positive controls.

Applicant's summary and conclusion

Conclusions:

The test item was considered to be non-mutagenic under the conditions of this test.

Executive summary:

The test method was designed to be compatible with the OECD Guidelines for Testing of Chemicals No. 471 "Bacterial Reverse Mutation Test".

Bisphenol F was cytotoxic for the bacteria at the highest concentration tested (500 µg/plate), especially without exogenous activation system. In this study, none of the results of the Ames test (+S9 or −S9) exceeded the critical value of 2.0 and all quotients ranged below 1.6, therefore the Ames test did not show any genotoxic potential of Bisphenol F, compared to the respective positive controls.