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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
data from handbook or collection of data
Justification for type of information:
Data is from peer reviewed publication

Data source

Reference
Reference Type:
publication
Title:
Salmonella Mutagenicity Tests: II. Results From The Testing Of 270 Chemicals
Author:
Mortelmans,K, Haworth,S, Lawlor,T, Speck,W, Tainer,B And Zeiger,E
Year:
1986
Bibliographic source:
Environ. Mutagen. 8(Suppl. 7):1 -119, 1986

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Principles of method if other than guideline:
Gene mutation toxicity study was performed for D-Carvone to evaluate its mutagenic nature
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material: D-Carvone- IUPAC name: (5S)-2-methyl-5-(prop-1-en-2-yl)cyclohex-2-en-1-one- Molecular formula: C10H14O- Molecular weight: 150.22 g/mol- Substance type: Organic
Specific details on test material used for the study:
- Name of test material: D-Carvone- IUPAC name: (5S)-2-methyl-5-(prop-1-en-2-yl)cyclohex-2-en-1-one- Molecular formula: C10H14O- Molecular weight: 150.22 g/mol- Substance type: Organic- Physical state: No data- Purity: 95.4%- Impurities (identity and concentrations): 4.6%

Method

Target gene:
Histidine
Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Remarks:
Lab 1 and Lab 2
Details on mammalian cell type (if applicable):
Not applicable
Additional strain / cell type characteristics:
not specified
Cytokinesis block (if used):
No data
Metabolic activation:
with and without
Metabolic activation system:
Male Sprague-Dawley rats and male Syrian hamsters were routinely used for the S9 preparation of the liver fractions
Test concentrations with justification for top dose:
0, 3.3, 10.0, 33.0, 100.0, 333.0 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO- Justification for choice of solvent/vehicle: The test chemical was soluble in DMSO
Controls
Untreated negative controls:
not specified
Negative solvent / vehicle controls:
yes
Remarks:
DMSO
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
9-aminoacridine
sodium azide
other: 4-nitro-o-phenylenediamine (TA98; -S9), 2-aminoanthracene (all strains, +S9)
Details on test system and experimental conditions:
METHOD OF APPLICATION: preincubationDURATION- Preincubation period: 20 mins- Exposure duration: 48 hr- Expression time (cells in growth medium): 48 hr- Selection time (if incubation with a selection agent): No data- Fixation time (start of exposure up to fixation or harvest of cells): No dataSELECTION AGENT (mutation assays): No dataSPINDLE INHIBITOR (cytogenetic assays): No dataSTAIN (for cytogenetic assays): No dataNUMBER OF REPLICATIONS: At least five dose levels of the chemicals were tested, with three plates per dose level.NUMBER OF CELLS EVALUATED: No dataDETERMINATION OF CYTOTOXICITY- Method: mitotic index; cloning efficiency; relative total growth; other: No dataOTHER EXAMINATIONS:- Determination of polyploidy: No data- Determination of endoreplication: No data- Other: No dataOTHER: No data
Rationale for test conditions:
No data
Evaluation criteria:
1) mutagenic response: a dose-related, reproducible increase in the number of revertants over background, even if the increase was less than twofold; 2) nomutagenic response: when no increase in the number of revertants was elicited by the chemical; 3) questionable response: when there was an absence of a clear-cut dose-related increase in revertants; when the dose-related increases in the number of revertants were not reproducible;or when the response was of insufficient magnitude to support a determination of mutagenicity
Statistics:
Mean and Standard error of mean

Results and discussion

Test results
Species / strain:
S. typhimurium, other: TA100, TA1535, TA1537, TA98
Remarks:
Lab 1 and Lab 2
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
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS- Effects of pH: No data - Effects of osmolality: No data- Evaporation from medium: No data- Water solubility: No data- Precipitation: No data- Other confounding effects: No dataRANGE-FINDING/SCREENING STUDIES: The chemical was initially tested with strain TA100 in the presence and the absence of the metabolic activation systems, over a wide dose range with an upper limit of 10 mg/plate, or less when solubility problems were encountered. Toxicity was evidenced by one or more of the following phenomena: appearance of his+ pinpoint colonies, reduced numbers of revertant colonies per plate, or thinning or absence of the bacterial lawn. Nontoxic chemicals were tested in the initial experiment up to the 10 mg/plate dose level, or to a level determined by their solubility. Toxic chemicals were tested up to a high dose which exhibited some degree of toxicity.COMPARISON WITH HISTORICAL CONTROL DATA: No data
Remarks on result:
other: No mutagenic potential

Any other information on results incl. tables

Table: Mutation data for the test chemical D- carvone

Dose (µg/plate)

TA100

NA

10% HLI

10% RLI

Mean

SEM

Mean

SEM

Mean

SEM

0

89

4.6

121

12.1

133

11.7

3.3

84

7.6

119

9.9

94

14.1

10.0

75

4.0

113

9.6

105

6.7

33.0

89

4.1

107

6.7

108

2.2

100.0

73

7.4

104

6.7

108

2.2

333.0

73

7.4

104

3.0

100

5.0

Positive control

48

9.0

78

8.4

65

11.3

 

Dose (µg/plate)

TA1535

NA

10% HLI

10% RLI

Mean

SEM

Mean

SEM

Mean

SEM

0

4

1.5

6

1.5

8

3.0

3.3

3

0.9

5

1.3

5

0.3

10.0

3

0.7

4

0.9

5

1.2

33.0

3

0.

3

0.7

4

1.5

100.0

2

1.0

5

0.3

3

0.6

333.0

1

0.6

4

0.6

3

2.2

Positive control

122

11.1

41

5.8

76

13.4

 

Dose (µg/plate)

TA1537

NA

10% HLI

10% RLI

Mean

SEM

Mean

SEM

Mean

SEM

0

3

0.7

6

0.6

6

1.3

3.3

3

0.6

5

0.7

5

1.2

10.0

2

0.0

5

1.2

8

 

33.0

3

1.2

2

0.7

7

1.7

100.0

3

1.0

4

0.7

8

2.7

333.0

3

0.6

6

1.2

6

0.9

Positive control

1041

154.2

261

12.0

211

24.1

 

Dose (µg/plate)

TA98

NA

10% HLI

10% RLI

Mean

SEM

Mean

SEM

Mean

SEM

0

14

1.9

25

1.2

21

3.8

3.3

16

2.8

23

5.0

20

3.2

10.0

15

0.3

23

1.2

15

3.2

33.0

11

1.2

25

2.5

22

0.9

100.0

13

0.9

23

2.0

24

1.5

333.0

10

2.1

17

4.0

22

2.0

Positive control

359

21.4

1969

56.9

973

201.3

Applicant's summary and conclusion

Conclusions:
D- carvone did not induce gene mutation in the Salmonella typhimurium strain TA100, TA1535, TA1537, TA98 both in the presence and absence of S9 metabolic activation system and hence the chemical is not likely to classify as a gene mutant in vitro.
Executive summary:

Gene mutation toxicity study was performed for D- carvone to evaluate its mutagenic nature. The study was performed as per the preincubation protocol using Salmonella typhimurium strain TA100, TA1535, TA1537, TA98 both in the presence and absence of S9 metabolic activation system at doses of0, 3.3, 10.0, 33.0, 100.0, 333.0 µg/plate. DMSO was used at the vehicle. The plates were incubated for 48 hrs after 20 mins preincubation before the evaluation of the revertant colonies could be made. D- carvone did notinduce gene mutation in theSalmonella typhimurium strain TA100, TA1535, TA1537, TA98 both in the presence and absence of S9 metabolic activation system and hence the chemical is not likely to classify as a gene mutant in vitro.