Etilefrine

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:

2000

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2000

Report date:

2000

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

Except the selection of the tester strains

Test guideline 2

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5265 (The Salmonella typhimurium Bacterial Reverse Mutation Test)

Deviations:

yes

Remarks:

Except the selection of the tester strains

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: EEC Directive 2000/32, L 136, Annex 4D, B13/14, dated June 08, 2000

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Etilefrine

EC Number:

211-910-4

EC Name:

Etilefrine

Cas Number:

709-55-7

Molecular formula:

C10H15NO2

IUPAC Name:

3-[2-(ethylamino)-1-hydroxyethyl]phenol

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

Name : Effortil-Rohbase
Batch No.: 310
Aggregate State at RT: solid
Colour: white
Purity: >98%
Analysis: meldting point
Certificate of Analysis: 19.07.2000
Sorage: in original container, at room temperature

Method

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 100

Additional strain / cell type characteristics:

other: his G46;rfa;uvrB;R-factor: base-pair substitutions

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 98

Additional strain / cell type characteristics:

other: his D3052; rfa; uvrB;R-factor: frame shift mutations

Metabolic activation:

with and without

Test concentrations with justification for top dose:

31.6; 100.0; 316.2; 1000.0; 2500.0 and 5000.0ug/plate

Controls

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

sodium azide

other: 4-Nitro-o-phenylene-diamine (4-NOPD); 2-aminoanthracene (2-AA)

Remarks:

The stability of the positive control substances in solution is unknown but a mutagenic response in the expected range is sufficient evidence of biological stability.

Details on test system and experimental conditions:

For the pre-incubation method 100 ul of the test item solution was preincubated with the tester strains (100ul) and sterile buffer or the metabolic activation system (500ul) for 60 minutes at 37°C prior to adding the overlay agar (2000ul) and pouring onto the surface of a minimal agar plate.
For each strain and dose level, including the controls, three plates were used.
After solidification the plates were inverted and incubated at 37°C for at least 48h in the dark.

Evaluation criteria:

The Mutation Factor is calculated by dividing the mean value of the revertant counts through the mean values of the solvent control (the exact and not the rounded values are used for calculation).

For positive controls solved in water:
due to a lack in the evaluation-software the mutation factor is correlated to the solvent control and not to the negative (water) control. This does not influence the validity of the data.

A test is considered acceptable if for each strain:
the bacteria demonstrate their typical responses to crystal violet and ampicillin
the control plates without S9 mix are within the following ranges (range of spontaneous reversion frequencies, historical control data range):
TA 98: 18-63
TA 100: 79-197
corresponding background growth an both negative control and test plates is observed.
the positive controls show a distinct enhancement over the control plate.

A test item is considered as mutagenic if:
- a dose-related increase in the number of revertants occurs and/or
- a reproducible biologically relevant positive response for at least one of the test points occurs
in at least one strain with or without metabolic activation.
According to the new OECD guidelines, the biological relevance of the results will be the criterion for the interpretation of results, a statistical evaluation of the results is not regarded as necessary.

A test item producing neither a dose related increase in the number of revertants nor a reproducible biologically relevant positive response at any of the test points is considered non-mutagenic in this system.
A biologically relevant increase is described as follows:
if in test strain TA 100 the number of reversions is at least twice as high
if in test strain TA 98 the number of reversions is at least three times higher
as compared to the spontaneous reversion rate.

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:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

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:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:

In conclusion, it can be stated that during the described mutagenicity test and under the experimental conditions reported, the test item did not cause gene mutations by base pair changes or frameshifts in the genome of the tester strains used.

Executive summary:

In order to investigate the potential of Effortil-Rohbase for its ability toinduce gene mutations the plate incorporation test (experiment I) and the pre­incubation test (experiment II) were perfoiuied with theSalmonellatyphimuriumstrains TA 98 and TA 100.

The test item was tested in two independent experiments at severalconcentrations. Each assay was conducted with and without metabolicactivation (S9 mix). The concentrations, including the controls, were tested intriplicate. The following concentrations of the test item were prepared andused in the experiments:

31.6; 100.0; 316.2; 1000.0; 2500.0 and 5000.0 µg/plate

Slight toxic effects indicated by a reduction of the background lawn of thetest item were observed in experiment II with metabolic activation with bothtester strains at 2500.0 and 5000.0 ug/plate. No toxic effects could beobserved in the first experiment and in the second experiment without metabolicactivation up to the highest investigated dose in both strains used.

No substantial increases in the revertant colony numbers of any of the twotest strains were detected at any dose level of the test item either with orwithout metabolic activation in both independently performed experiments.

As positive controls reference mutagens were tested in parallel to the testitem. They showed a distinct increase of induced revertant colonies