3,4-dihydro-2H-pyran

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1999

Report date:

1999

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

3,4-dihydro-2H-pyran

EC Number:

203-810-4

EC Name:

3,4-dihydro-2H-pyran

Cas Number:

110-87-2

Molecular formula:

C5H8O

IUPAC Name:

3,4-dihydro-2H-pyran

Details on test material:

TS-Freetext:
3,4-Dihydropyran, purity 99%.

Method

Species / strain

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 from male rat livers

Test concentrations with justification for top dose:

20 - 5000 ug/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

9-aminoacridine

N-ethyl-N-nitro-N-nitrosoguanidine

other: N-methyl-N`-nitro-N-nitrosoguanidine, 4-nitro-o-phenylendiamine

Details on test system and experimental conditions:

Standard plate test
• Salmonella typhimurium
The experimental procedure of the standard plate test (plate incorporation method) is based on the method of Ames et al. Test tubes containing 2-ml portions of soft agar (overlay agar), which consists of 100 ml agar (0.6% agar + 0.6% NaCl) and 10 ml amino acid solution (minimal amino acid solution for the determination of mutants: 0.5 mM histidine + 0.5 mM biotin) are kept in a water bath at 45°C, and the remaining components are added in the following order:
0.1 ml test solution or vehicle
0.1 ml fresh bacterial culture
0.5 ml S-9 mix (in tests with metabolic activation)
or
0.5 ml phosphate buffer (in tests without metabolic activation)
After mixing, the samples are poured onto Vogel-Bonner agar plates (minimal glucose agar plates) within approx. 30 seconds.
Composition of the minimal glucose agar:
980 ml aqua dest.
20 ml Vogel-Bonner E medium
15 g Difco bacto agar
20 g D-glucose, monohydrate.
After incubation at 37°C for 48 - 72 hours in the dark, the bacterial colonies (his• revertants) are counted.
• Escherichia coli
The experimental procedure is based on the method of Ames et al. Test tubes containing 2-ml portions of soft agar (overlay agar), which consists of 100 ml agar (0.6% agar + 0.6% NaCl) and 10 ml amino acid solution (minimal amino acid solution for the determination of mutants: 0.5 mM tryptophan) are kept in a water bath at 45°C, and the remaining components are added in the following order:
0.1 ml test solution or vehicle
0.1 ml fresh bacterial culture
0.5 ml S-9 mix (in tests with metabolic activation)
or
0.5 ml phosphate buffer (in tests without metabolic activation)
After mixing, the samples are poured onto minimal agar plates within approx. 30 seconds.
Composition of the minimal agar:
The composition of the minimal agar (SA1 selective agar) is based on the description of Green, M.H.L. and Muriel, W.J., with the exception of solution E (tryptophan solution), which has previously been added to the soft agar:
300 ml solution B (agar)
100 ml solution A (saline solution)
8 ml solution C (glucose solution)
10 ml solution D (casein solution)
After incubation at 37°C for 48 - 72 hours in the dark, the bacterial colonies (trp+ revertants) are counted.
Preincubation test
The experimental procedure is based on the method described by Yahagi et al. and Matsushima et al.
0.1 ml test solution or vehicle, 0.1 ml bacterial suspension and 0.5 ml S-9 mix are incubated at 37°C for the duration of about 20 minutes using a shaker. Subsequently, 2 ml of soft agar is added and, after mixing, the samples are poured onto the agar plates within approx. 30 seconds. After incubation at 37°C for 48 - 72 hours in the dark, the bacterial colonies are counted.

Results and discussion

Test results

Species / strain:

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

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

other: from about 500 - 2500 ug/plate upwards

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: all strains/cell types tested

Any other information on results incl. tables

Solubility: No prediction of the test substance was found.

Toxicity: A slight decrease in the number of revertats was occasionally observed in the PIT depending on the strain and test conditions from about 500 µg - 2500 µg/plate onward.

Mutagenicity: An increase in the number of his+ or trp* revertants was not observed in the standard plate test or in the preincubation test either without S-9 mix or after the addition of a metabolizing system.

Applicant's summary and conclusion

Conclusions:

According to the results of the present study, the test substance 3,4-dihydropyran is not mutagenic in the Salmonella typhimurium/Escherichia coli reverse mutation assay (Ames test) under the experimental conditions chosen.

Executive summary:

The substance 3,4-Dihydropyran was tested for its mutagenic potential based on the ability to induce point mutations in selected loci of several bacterial strains, i.e. Salmonella typhimurium and Escherichia coli, in a reverse mutation assay. An increase in the number of revertants was not observed in the standard plate test or in the preincubation test either without S-9 mix or after the addition of a metabolizing system.