2-octyldodecyl 12-[(1-oxooctadecyl)oxy]octadecanoate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1994

Report date:

1994

Reference 2

Reference Type:

secondary source

Title:

Unnamed

Year:

2001

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

adopted 1997

Deviations:

yes

Remarks:

S. typhimurium strain TA 102 or an E. coli strain not included, only 2-aminoacridine used as the positive control with metabolic activation and the S-9 mix was not characterised with a mutagen requiring metabolic activation

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Method

Target gene:

his operon

Species / strain

Species / strain / cell type:

S. typhimurium, other: TA 1535, TA 1537, TA 98, TA 100 and TA 1538

Metabolic activation:

with and without

Metabolic activation system:

cofactor supplemented post-mitochondrial fraction (S9 mix), prepared from the livers of rats treated with Aroclor 1254

Test concentrations with justification for top dose:

Experiment I and II: 1.0, 5.0, 10, 50 and 100 µL/plate (all strains), with and without metabolic activation

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: acetone
- Justification for choice of solvent/vehicle: the test substance was miscible to 1mL/mL in acetone and corn oil. The stock solution of the test substance was prepared in acetone, therefore acetone was selected as the vehicle

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

acetone

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: sodium azide (1 µg/plate, -S9, TA 1535, TA 100); 9-aminoacridine (50 μg/plate, -S9, TA 1537); 2-nitrofluorene (5 μg/plate, -S9, TA 98, TA 1538); 2-aminoanthracene (1.25 μg/plate, +S9,TA 1535, TA 1537, TA 98, TA 100, TA 1538)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

NUMBER OF REPLICATIONS: triplicates in each of two independent experiments

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth (thinning of bacterial background lawn); reduction in the number of revertant colonies per plate

OTHER:
In a preliminary range-finding test, the cytogenicity of the test substance was assessed. TA 100 was treated with 0.05, 0.1, 0.5, 1.0, 5.0, 10, 50 and 100 µL/plate, with and without metabolic activation. The genotypes of the tester strains were confirmed by testing for histidine requirement and rfa-mutation, and TA and TA 100 were confirmed positive for the pKM101 plasmid. The viabilty of the tester strain was confirmed. The sterility of the S9-mix was checked before the beginning and the end of each experiment and was found to be satisfactory.

Evaluation criteria:

Evaluation criteria:
A response is considered positive if at least one strain has a dose that produces a mean reversion frequency that is 2 times or more greater than the mean reversion frequency of the corresponding solvent control plates and the response is dose dependent. The degree of toxicity will be considered in relation to the results. A response is considered equivocal if it does not fulfill the criteria of either negative or positive response and/or the study director does not consider the reponse to be either positive or negative.

Statistics:

Mean values and standard deviations were calculated for the number of revertants per plate.

Results and discussion

Test results

Species / strain:

S. typhimurium, other: TA 1535, TA 1537, TA 98, TA 100 and TA 1538

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Water solubility: the test substance was soluble up to 1 mL/mL in acetone and corn oil
- Precipitation: slight precipitation was observed in the range-finding assay from 50 µL/plate (TA 100) with and without metabolic activation and in the main assays with and withour metabiolic activation.

RANGE-FINDING/SCREENING STUDIES: cell viability was comparable between control and treated strains up to the highest concentration level of 100 µL/plate. Slight precipitation was observed in the range-finding assay from 50 µL/plate (TA 100) with and without metabolic activation.

Any other information on results incl. tables

Table 1: Experiment 1

EXPERIMENT 1 (plate incorporation test)
S9-Mix	Without
Test item (µL/plate)	TA 98	TA 100	TA 1535	TA 1537	TA 1538
NC	26 ± 5.9	80 ± 4.0	21 ± 4.0	12 ± 3.6	10 ± .6
1.0	24 ± 2.3	80 ± 3.8	21 ± 1.5	13 ± 3.1	10 ± 3.8
5.0	24 ± 1.5	78 ± 1.5	19 ± 3.8	11 ± 2.9	9 ± 4.0
10	29 ± 5.9	69 ± 2.0	16 ± 3.5	11 ± 3.1	10 ± 2.6
50	27 ± 3.6*	85 ± 4.9*	16 ± 0.6*	9 ± 2.6*	10 ± 2.1*
100	23 ± 3.5*	82 ± 12.8*	20 ± 0.6*	9 ± 3.0*	9 ± 4.0*
2-NF	775 ± 71.5	-	-	-	857 ± 60.2
SA	-	476 ± 4.7	456 ± 16.7	-	-
9-AA	-	-	-	86 ± 10.8	-
S9-Mix	With
Test item (µL/plate)	TA 98	TA 100	TA 1535	TA 1537	TA 1538
NC	26 ± 6.0	57 ± 4.0	11 ± 1.7	13 ± 1.2	14 ± 1.7
1.0	31 ± 6.7	61 ± 0.6	13 ± 1.5	13 ± 2.3	14 ± 1.5
5.0	34 ± 1.2	75 ± 6.0	14 ± 4.0	11 ± 3.1	16 ± 7.2
10	32 ± 2.5	63 ± 4.4	16 ± 1.2	11 ± 2.6	11 ± 5.0
50	29 ± 4.0*	67 ± 2.0*	15 ± 0.6*	10 ± 3.6*	11 ± 4.9*
100	27 ± 3.6*	61 ± 1.7*	15 ± 1.5*	11 ± 2.1*	12 ± 0.6*
2AA	665 ± 43.9	716 ± 68.0	366 ± 1.5	138 ± 1.0	720 ± 114.1
*slight precipitate NC = Vehicle Control, acetone 2-NF: 2-nitrofluorene SA: sodium azide 9AA:9-aminoacridine 2AA: 2-aminoanthracene

 

Table 2: Experiment 2

EXPERIMENT 2 (plate incorporation test)
S9-Mix	Without
Test item (µL/plate)	TA 98	TA 100	TA 1535	TA 1537	TA 1538
NC	27 ± 2.6	66 ± 2.1	20 ± 0.6	11 ± 3.1	11 ± 1.2
1.0	24 ± 1.5	70 ± 2.6	19 ± 2.9	12 ± 2.9	11 ± 2.1
5.0	25 ± 3.1	71 ± 2.5	19 ± 2.1	13 ± 5.5	12 ± 2.0
10	28 ± 3.0	56 ± 2.1	19 ± 4.6	12 ± 2.5	12 ± 2.5
50	26 ± 4.7*	63 ± 5.5*	23 ± 1.2*	11 ± 1.2*	11 ± 1.7*
100	21 ± 3.2*	61 ± 3.8*	21 ± 2.6*	10 ± 2.5*	9 ± 0.6*
2-NF	781 ± 73.2	-	-	-	919 ± 97.6
SA	-	486 ± 31.3	462 ± 14.6	-	-
9-AA	-	-	-	100 ± 6.0	-
S9-Mix	With
Test item (µL/plate)	TA 98	TA 100	TA 1535	TA 1537	TA 1538
NC	33 ± 6.5	57 ± 6.6	17 ± 1.5	14 ± 1.5	13 ± 1.0
1.0	36 ± 4.0	57 ± 3.1	16 ± 5.5	12 ± 2.5	12 ± 0.6
5.0	36 ± 2.6	57 ± 8.9	14 ± 4.0	12 ± 2.0	12 ± 2.3
10	33 ± 3.1	58 ± 11.7	14 ± 1.2	12 ± 0.6	12 ± 3.0
50	37 ± 3.8*	58 ± 10.1*	15 ± 1.0*	10 ± 2.5*	15 ± 4.9*
100	33 ± 7.0*	60 ± 9.0*	12 ± 2.1*	13 ± 3.2*	13 ± 0.6*
2AA	699 ± 50.8	802 ± 97.2	344 ± 5.9	122 ± 8.1	673 ± 68.5
*slight precipitate NC = Vehicle Control, acetone 2-NF: 2-nitrofluorene SA: sodium azide 9AA:9-aminoacridine 2AA: 2-aminoanthracene

 

 

 

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative