Uridine, 2'-deoxy-5-ethyl-, 3',5'-bis(4-chlorobenzoate)

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:

December 12th 2000 - January 8th 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2001

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EEC Council Directive 2000/32, Annex 4D.

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: ICH S2A Genotoxicity: Specific Aspect of Regulatory Tests

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Uridine, 2'-deoxy-5-ethyl-, 3',5'-bis(4-chlorobenzoate)

EC Number:

607-566-3

Cas Number:

25137-84-2

Molecular formula:

C25H22Cl2N2O7

IUPAC Name:

Uridine, 2'-deoxy-5-ethyl-, 3',5'-bis(4-chlorobenzoate)

Test material form:

solid: particulate/powder

Specific details on test material used for the study:

- Name of test material (as cited in study report): 2’DEOXY-5-ETHYL-URIDINE, 3’5’-BIS(4-CHLOROBENZOATE)
- Physical state: Solid (whitish powder)
- Lot/batch No.: 2/98-G
- Expiration date of the lot/batch: 30th November 2003
- Storage: At room temperature

Method

Target gene:

TA1535, TA1537, TA98, TA100 and TA102

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Metabolic activation:

with and without

Metabolic activation system:

S9

Test concentrations with justification for top dose:

EXPERIMENT I: 5000, 2500, 1250, 625 and 313 µg/plate
EXPERIMENT II: 5000, 2500, 1250, 625 and 313 µg/plate

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

Positive controls:

yes

Positive control substance:

9-aminoacridine

2-nitrofluorene

sodium azide

cumene hydroperoxide

other: 2-Aminoanthracene

Remarks:

sterile distilled water

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation); preincubation

DURATION
- Preincubation period: 30 minutes at 37 °C
- Exposure duration: 72 hours at 37 °C

DETERMINATION OF CYTOTOXICITY
- Method: Toxicity was assessed on the basis of a decline in the number of spontaneous revertants, a thinning of the background lawn or a microcolony formation. The scoring was effected by counting the number of revertant colonies on each plate

Evaluation criteria:

For the test item to he considered mutagenic, two-fold (or more) increases in mean revertant numbers must be observed at two consecutive dose-levels or at the highest practicable dose-level only. In addition, there must be evidence of a dose-response relationship showing increasing numbers of mutant colonies with increasing dose levels. The effect must be reproduced in an independent experiment

Statistics:

A statistical evaluation of the results is not regarded as necessary

Results and discussion

Test results

Key result

Species / strain:

S. typhimurium, other: TA1535, TA1537, TA98, TA100 and TA102

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Remarks on result:

other: EXPERIMENT I: 5000, 2500, 1250, 625 and 313 µg/plate

Applicant's summary and conclusion

Conclusions:

It is concluded that the test item 2’DEOXY-5-ETHYL-URIDINE, 3’5’-BIS(4-CHLOROBENZOATE) does not induce reverse mutation in Salmonella typhimurium under the reported experimental conditions.

Executive summary:

The test item 2’DEOXY-5-ETHYL-URIDINE, 3’5’-BIS(4-CHLOROBENZOATE) was examined for mutagenic activity by assaying for reverse mutation to histidine prototrophy in the prokaryotic organism Salmonella typhimurium.

The five tester strains TA1535, TA1537, TA98, TA100 and TA102 were used.

Experiments were performed both in the absence and presence of metabolic activation, using liver S9 fraction from rats pre-treated with phenobarbitone and betanaphthoflavone.

Test item solutions were prepared using dimethylsulphoxide.

In the toxicity test, the test item was assayed at a maximum dose-level of 5000 µg/plate and at four lower dose-levels spaced at approximately half-log intervals: 1580, 500, 158 and 50.0 µg/plate. No signs of toxicity were observed at any dose-level tested, in any tester strain, in the absence or presence of S9 metabolic activation.

Two main experiments were performed.

In Main Assay I, using the plate incorporation method, the test item was assayed at a maximum dose-level of 5000 µg/plate and at four lower dose-levels, separated by two-fold dilutions: 2500, 1250, 625 and 313 µg/plate.

As no increases in revertant numbers were observed, all treatments of Main Assay II included a pre-incubation step and used the same dose-range employed in Main Assay I. Moderate toxicity was observed at higher dose-levels with all tester strains both in the absence and presence of S9 metabolism.

Precipitation of the test item was observed at higher dose-levels in both experiments.

The test item did not induce two-fold increases in the number of revertant colonies in the plate incorporation or pre-incubation assay, at any dose-level, in any tester strain, in the absence or presence of S9 metabolism.

It is concluded that the test item 2’DEOXY-5-ETHYL-URIDINE, 3’5’-BIS(4-CHLOROBENZOATE) does not induce reverse mutation in Salmonella typhimurium under the reported experimental conditions.