Diphenyl sulphone

Genetic toxicity: in vitro

Administrative data

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:

1989

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Meets scientific standards and GLP study, but acceptable restrictions (stability of test substance not analysed).

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1989

Report date:

1989

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

the stability and concentration of the test substance in the vehicle used were not determined by analysis.

Principles of method if other than guideline:

Minor modifications

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Diphenyl sulphone

EC Number:

204-853-1

EC Name:

Diphenyl sulphone

Cas Number:

127-63-9

Molecular formula:

C12H10O2S

IUPAC Name:

(benzenesulfonyl)benzene

Test material form:

solid: crystalline

Details on test material:

- Name of test material (as cited in study report): diphenyl sulphone
- Substance type: monoconstituent
- Physical state: white crystalline solid
- Analytical purity: assumed purity 100%
- Impurities (identity and concentrations): not applicable
- Composition of test material, percentage of components: assumed purity 100%
- Isomers composition: not applicable
- Purity test date: see confidential details
- Lot/batch No.: see confidential details
- Expiration date of the lot/batch: see confidential details
- Stability under test conditions: stable under normal storage and test conditions according the Sponsor
- Storage condition of test material: at ambient temperature in the dark until required
- Other: not applicable

Method

Target gene:

Hisitdine

Species / strain

Species / strain / cell type:

S. typhimurium, other: TA 1535, TA1537, TA1538, TA98, TA100

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

S9

Test concentrations with justification for top dose:

Experiment 1 (dose range finding): 1.6; 8.0; 40; 200; 1000 and 5000µg/plate (TA100 +S9 and –S9)
Experiment 2: 1.6; 8.0; 40; 200; 1000 and 5000µg/plate (TA1535, TA1537, TA 1538, TA98, TA100 all +S9)
Experiment 3: 1.6; 8.0; 40; 200; 1000 and 5000µg/plate (TA1535, TA1537, TA 1538, TA98, TA100 all –S9)
Experiment 4: 1.6; 8.0; 40; 200; 1000 and 5000µg/plate (TA1535, TA1537, TA 1538, TA98, TA100 all +S9)
Experiment 5: 1.6; 8.0; 40; 200; 1000 and 5000µg/plate (TA1535, TA1537, TA 1538, TA98, TA100 all –S9)

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO (dimethylsulphoxide) (BDH Ltd, Poole, Dorset, UK: CTL Reference number Y00876/001/105)
= Solvent for the positive control substances , for the test compound and the negative control.

Controls

Controls 1

Untreated negative controls:

yes

Remarks:

Untreated

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

Positive controls:

yes

Remarks:

+S9:2-Aminoanthracene. -S9:N-Methyl-N'-nitro-N-notrosoguanidine

Positive control substance:

other:

Remarks:

Experiment 1 (dose range finding) TA100

Controls 2

Untreated negative controls:

yes

Remarks:

Untreated

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

Positive controls:

yes

Remarks:

+S9 TA1535 and TA1537: 2-Aminoanthracene. +S9 TA1538 , TA98 and TA100: 2-Aminoanthracene

Positive control substance:

other:

Remarks:

Experiment 2

Controls 3

Untreated negative controls:

yes

Remarks:

Untreated

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

Positive controls:

yes

Remarks:

all -S9:TA1535: N-Methyl-N’-nitro-N-nitrosoguanidine.TA37: Acridine Mutagen ICR191.TA1538: 4-Nitro-O-Phenylenediamine. TA98: Daunomycin HCl (DR): . TA100: N-Methyl-N’-nitro-N-nitrosoguanidine

Positive control substance:

other:

Remarks:

Experiment 3

Controls 4

Untreated negative controls:

yes

Remarks:

Untreated

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

Positive controls:

yes

Remarks:

+S9: 2-Aminoanthracene: TA 1535, TA 1537, TA1538, TA98 and TA100.

Positive control substance:

other:

Remarks:

Experiment 4

Controls 5

Positive controls:

yes

Remarks:

All -S9:TA1535: N-Methyl-N’-nitro-N-nitrosoguanidine (MNNG); TA1537: Acridine Mutagen ICR191; TA1538: 4-Nitro-O-Phenylenediamine; TA98: Daunomycin HCl (DR); TA100: N-Methyl-N’-nitro-N-nitrosoguanidine (MNNG)

Positive control substance:

other:

Remarks:

Experiment 5

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation); method described by Maron and Ames (1983).

DURATION
- Preincubation period:not applicable (plate incorporation)
- Exposure duration:66 hours (incubation period)
- Expression time (cells in growth medium): not applicable
- Selection time (if incubation with a selection agent): not applicable
- Fixation time (start of exposure up to fixation or harvest of cells): not applicable

SELECTION AGENT (mutation assays): Vogel Bonner minimal medium containing 1.5% w/v agar and 2% w/v glucose (Gibco-Europe LTD) present in petri-dishes; 0.5 mM Histidine/0.5 MB Biotin was prepared and fiter-sterilized; top agar consisted of 0.6% w/v agar and 0,5% w/W NaCl in dionised water molten at 50-53°C was spiked wiht the histidine and biotin solution (10 ML solution in 100 mL agar).

NUMBER OF REPLICATIONS: 3

NUMBER OF CELLS EVALUATED: not applicable

DETERMINATION OF CYTOTOXICITY
- Method:other: number

Evaluation criteria:

Test data from individual experiments are considered valid if:
a) the concurrent solvent control data are acceptable;
b) the positive control data show equivocal positive responses;
c) at least the lowest test compound dose shows no evidence of toxicity, and at least three test doses show no significant overt toxicity (ie significant loss of background growth and/or reductions in colony numbers).
Failure of one or more tester strain/S9 combinations does not invalidate the data for the remainder of a concurrent experiment.
A positive response in a (valid) individual experiment is achieved when one or both of the following criteria are met:
a) a statistically significant dose-related increase in the number of revertant colonies is obtained;
b) a two-fold or greater increase in the mean number of revertant colonies (over that observed for the concurrent solvent control plates) which is statistically significant, is observed at at least one dose level.
A negative result in a (valid) experiment is achieved when:
a) There is no statistically significant dose-related increase in the mean number of revertant colonies per plate observed for the test compound; and
b) in the absence of any such dose response, no increase in colony numbers is observed (at any test dose) which exceeds 2x the concurrent solvent control.
For a positive response in an individual experiment to be considered indicative of an unequivocal positive, ie mutagenic, result for that strain/S9 combination, then the observed effect(s) must be consistently reproducible.

Statistics:

The assessment of statistical significance was carried out using a one-tailed Student’s t-test (Ehrenberg 1984) . The corresponding probability for each dose level was derived by computer using the the appropriate degrees of freedom. Values of p<0.01 were treated as significant, with values of 0.01

Results and discussion

Test results

Test results 1

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

Positive controls validity:

valid

Test results 2

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

Positive controls validity:

valid

Test results 3

Key result

Species / strain:

S. typhimurium TA 1538

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 4

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

Positive controls validity:

valid

Test results 5

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:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: not applicable
- Effects of osmolality: not applicable
- Evaporation from medium: not applicable
- Water solubility: not applicable
- Precipitation: The precipitation observed at the top dose tested in each experiment shows that the compound was tested to an appropriate maximum dose in each case.
- Other confounding effects:

RANGE-FINDING/SCREENING STUDIES: In the initial dose-range finding study, the compound showed no significant toxicity in strain TA100, either in the presence or absence of S9. The same dose range was therefore selected for the main experimental work.

COMPARISON WITH HISTORICAL CONTROL DATA: not applicable

ADDITIONAL INFORMATION ON CYTOTOXICITY: not applicable

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:

Under the conditions of this assay, diphenyl sulphone gave an unequivocal negative, i.e. non-mutagenic, response in both the presence and absence of a metabolizing system (S9), in all five strains of Salmonella used (TA1535, TA1537, TA1538, TA98 and TA100).

Executive summary:

Diphenyl sulphone has been evaluated in the Salmonella mutagenicity assay of Maron and Ames (1983), following a protocol complying with OECD (1983) Guideline Number 471.

In both the presence and absence of a metabolizing system (S9), the compound did not induce any significant reproducible increases in the observed numbers of revertant colonies in any of the five tester strains used (TA1535, TA1537, TA1538, TA98 and TA100). In each experiment, the positive control chemicals gave the expected responses, indicating that the assay system was working correctly.

Under the conditions of this assay, diphenyl sulphone therefore gave an unequivocal negative, ie non-mutagenic, response in both the presence and absence of an auxiliary metabolising system (S9),when tested to a maximum dose of 5000µg/plate, at which concentration significant precipitation of the compound was observed on the test plates.