1,2-epoxybutane

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1989-07-14 to 1989-07-20

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

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:

only 4 strains used

GLP compliance:

no

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

1,2-epoxybutane

EC Number:

203-438-2

EC Name:

1,2-epoxybutane

Cas Number:

106-88-7

Molecular formula:

C4H8O

IUPAC Name:

oxolane

Test material form:

liquid

Method

Target gene:

The Salmonella typhimurium histidine (his) reversion system measures his- → his+ reversions.

Species / strain

Species / strain / cell type:

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

Metabolic activation:

with and without

Metabolic activation system:

Type and composition of metabolic activation system:
- source of S9: rat liver S9 mix
- method of preparation of S9 mix: 5 male Sprague-Dawley rats (200 - 300 g) receive a single intraperitoneal injection of 500 mg Aroclor 1254 (as a 20% solution in peanut oil - w/v) per kg body weight 5 days before sacrifice. During this time the animals are housed in Makrolon cages in air-conditioned rooms. The day/night rhythm is 12 hours (light period from 6.00 - 18.00 hours and dark period from 18.00 - 6.00 hours). Standardized pelleted feed and tap water from bottles are available ad libitum. 5 days after administration the rats are sacrificed, and the livers are prepared (all preparation steps for obtaining the liver microsome enzymes are carried out using sterile solvents and glassware at a temperature of +4°C). The livers are weighed and washed in an equivalent volume of a 150 mM KCl solution (1 mL = 1 g wet liver), then cut into small pieces and homogenized in three volumes of KCl solution. After centrifugation of the homogenate at 9000 x g for 10 minutes at +4°C, 5-ml portions of the supernatant (so-called S-9 fraction) are quickly deep-frozen in dry ice and stored at -70°C to -80°C for 2 months at the most. The S-9 mix is prepared freshly prior to each experiment. For this purpose, a sufficient amount of S-9 fraction is thawed at room temperature and 3 volumes of S-9 fraction are mixed with 7 volumes of S-9 supplement (cofactors). This preparation was kept on ice until used.
- concentration or volume of S9 mix and S9 in the final culture medium: 0.5 mL
- quality controls of S9: no data

Test concentrations with justification for top dose:

20, 100, 500, 2500 and 5000 µg/plate (1st experiment with and without S9 mix)
100, 500, 2500, 5000 and 7500 ug/plate (2nd experiment with and without S9 mix)

Vehicle / solvent:

- Vehicle/solvent used: DMSO

Controls

Controls 1

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

100 µg, without S9 mix (TA 1537)

Controls 2

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 4-nitro-o-phenylenediamine

Remarks:

10 µg, without S9 mix (TA 98)

Controls 3

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: N-methyl-N'-nitro-N-nitroso- guanidine (MNNG)

Remarks:

5 µg, without S9 mix (TA 100, TA 1535)

Controls 4

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene

Remarks:

10 µg, with S-9 mix (TA 100, TA 98, TA 1537, TA 1535)

Details on test system and experimental conditions:

NUMBER OF REPLICATIONS:
- Number of cultures per concentration: triplicate
- Number of independent experiments: 2

METHOD OF TREATMENT/ EXPOSURE:
- Test substance added in agar (plate incorporation)

TREATMENT AND HARVEST SCHEDULE:
- Exposure duration/duration of treatment: 48 hours at 37°C

METHODS FOR MEASUREMENT OF CYTOTOXICITY
- Method: reduced background growth

Evaluation criteria:

In general, a substance to be characterized as positive in the Ames test has to fulfill the following requirements:
- doubling of the spontaneous mutation rate (control)
- dose-response relationship
- reproducibility of the results.

Statistics:

none

Results and discussion

Test results

Test results 1

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

not determined

Cytotoxicity / choice of top concentrations:

not determined

Vehicle controls validity:

not examined

Untreated negative controls validity:

not examined

True negative controls validity:

not examined

Positive controls validity:

not examined

Test results 2

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

True negative controls validity:

not applicable

Positive controls validity:

valid

Test results 3

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

True negative controls validity:

not applicable

Positive controls validity:

valid

Test results 4

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

True negative controls validity:

not applicable

Positive controls validity:

valid

Test results 5

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

True negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation and time of the determination: no data

STUDY RESULTS
Tests without S-9 mix:
TA 1535: Weakly positive reaction at 2500 µg/plate (factor 3.3) and at 5000 µg/plate (factor 3.4).
TA 100: Slight increase in the number of revertant colonies at 2500 µg/plate (factor 1.7 - 1.9) - 7500 µg/plate (factor 3.1).
TA 1537: No increase in the number of revertants.
TA 98: No increase in the number of revertants.

Tests with S-9 mix:
TA 1535: Weakly positive reaction at 2500 µg - 5000 µg/plate (factor 1.9 - 3.3)
TA 100: Slight increase in the mutation rate at 5000 µg and 7500 µg/plate (factor 1.8 - 2.6).
TA 1537: No increase in the number of revertants.
TA 98: No increase in the number of revertants.

-Concurrent vehicle negative and positive control data: see attached tables

- Signs of toxicity: No bacteriotoxic effect (reduced his- background growth) was observed.
- Individual plate counts: see attached tables

HISTORICAL CONTROL DATA: no data