Isobutyl laurate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Study period:

24 - 27 Jan 2002

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP guideline study with acceptable restrictions analytical purity of the test substance not specified).

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2002

Report date:

2002

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

analytical purity of test substance not specified;

GLP compliance:

yes (incl. QA statement)

Remarks:

The Department of Health of the Government of the United Kingdom

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

441-620-5

EC Name:

-

Cas Number:

163961-32-8

Molecular formula:

C20H40O2 (C16iso) C22H44O2 (C18iso) C22H42O2 (C18:1iso)

IUPAC Name:

Fatty acids, C16-18 and C18-unsatd., branched and linear, butyl esters

Method

Target gene:

his operon

Species / strain

Species / strain 1

Species / strain / cell type:

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

Additional strain / cell type characteristics:

not specified

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 102

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

S9-Mix

Test concentrations with justification for top dose:

Experiment I+II:
- 50, 150, 500, 1500 and 5000 µg/plate (with and without metabolic activation)

Vehicle / solvent:

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

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

Acetone

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: N -etþl-N'-nitro -N-nitroso guanidine (ENNG, 3µg/plate (TA100), 5µg/plate (TA1535)); 9-Aminoacridine (9AA, 80µg/plate (TA1537)); Mitomycin C (MMC, 0.5µg/plate (TA102); 4-Nitroquinoline- 1 -oxide (4NQO, 0.2 µg/plate (TA98))

Remarks:

without S9-mix

Controls 2

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

Acetone

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-Aminoanthracene (2-AA, 1µg/plate, TA 100; 2µg/plate, TA 1535 and TA 1537); 1,8 Dihydroxyanthraquinone (DAN, 10µg/plate, TA 102); Benzo(a)pyren (BP, 5µg/plate, TA 98)

Remarks:

with S9-mix

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 37 °C for approximately 48 hours

NUMBER OF REPLICATIONS: triplicates

DETERMINATION OF CYTOTOXICITY
- Method: Reduction of growth of bacterial background lawn

The frequency of revertant colonies was assessed using a Domino colony counter.

Evaluation criteria:

The reverse mutation assay may be considered valid if the following criteria are met:
- All tester strain cultures exhibit a characteristic number of spontaneous revertants per plate in the vehicle and untreated controls compared to historical controls.
- The appropriate characteristics for each tester strain have been confirmed.
- All tester strain cultures should be in the approximate range of bacteria per ml.
- Each mean positive control value should be at least two times the respective vehicle control value for each strain.
- There should be a minimum of four non-toxic test material dose levels and no evidence of excessive contamination.

The test material may be considered positive in this test system if the following criteria are met:
- The test material should have induced a reproducible, dose-related and statistically signifrcant increase in the revertant count in at least one strain of
bacteria.

Statistics:

Mean and standard deviation were calculated. Significance was tested using Dunnett's method of linear regression (Kirkland DJ (Ed) (1989) Statistical Evaluation of Mutagenicity Test Data. UKEMS Subcommittee on Guidelines for Mutagenicity Testing, Report - Part III, Cambridge University Press.)

Results and discussion

Test results

Test results 1

Species / strain:

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

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

Test results 2

Species / strain:

S. typhimurium TA 102

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
The test material caused no visible reduction in the growth of the bacterial background lawn at any dose level.
The test material was, therefore, tested up to the maximum recommended dose level of 5000 µg/plate.
A globular, oily, opaque precipitate was observed at 5000 µg/plate, this did not prevent the scoring of revertant colonies.

RANGE-FINDING/SCREENING STUDIES:
The test material was non-toxic to the strain of Salmonella used (TA100). The test material formulation and the S9-mix used in this experiment were both shown to be effectively sterile.

COMPARISON WITH HISTORICAL CONTROL DATA:
A history profile of vehicle and positive control values was presented.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
In a preliminary toxicity study the test material was non-toxic to the Salmonella strain TA 100.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Table 1: Results from Experiment 1:

With or without S9-Mix	Test substance concentration (μg/plate)	Mean number of revertant colonies per plate (average of 3 plates)
		TA 100	TA1535	TA102	TA98	TA1537
-	0	90	20	317	23	18
-	50	94	19	333	22	17
-	150	96	20	315	21	14
-	500	96	24	343	23	19
-	1500	94	26	357	27	20
-	5000	101	24	349	22	15
Positive controls - S9	Name	ENNG	ENNG	MMC	4NQO	9AA
	Concentrations (μg/plate)	3.0	5.0	0.5	02	80
	Number of colonies/plate	439	464	740	96	1510
+	0	98	16	380	37	25
+	50	102	15	375	42	25
+	150	101	15	370	40	27
+	500	110	14	355	44	17
+	1500	104	17	358	44	24
+	5000	103	15	358	38	30
Positive controls + S9	Name	2AA	2AA	DAN	BP	2AA
	Concentrations (μg/plate)	1.0	2.0	10	5	2.0
	Number of colonies/plate	2112	269	06	232	217

 

Table 2: Results from Experiment 2:

With or without S9-Mix	Test substance concentration (μg/plate)	Mean number of revertant colonies per plate (average of 3 plates)
		TA 100	TA1535	TA102	TA98	TA1537
-	0	76	21	274	26	11
-	50	68	23	270	25	8
-	150	81	24	300	21	9
-	500	80	25	281	23	10
-	1500	85	25	290	24	12
-	5000	76	26	280	24	14
Positive controls - S9	Name	ENNG	ENNG	MMC	4NQO	9AA
	Concentrations (μg/plate)	3.0	5.0	0.5	02	80
	Number of colonies/plate	679	933	1167	188	2357
+	0	78	14	373	39	19
+	50	77	14	385	46	27
+	150	86	15	401	45	21
+	500	82	14	382	42	19
+	1500	85	16	400	44	20
+	5000	91	19	379	39	16
Positive controls + S9	Name	2AA	2AA	DAN	BP	2AA
	Concentrations (μg/plate)	1.0	2.0	10	5	2.0
	Number of colonies/plate	3159	313	1021	312	222

2AA: 2-Aminoanthracene

BP: Benzo(a)pyrene

DAN: 1,8 Dihydroxyanthraquinone

ENNG: N-ethyl-N'-nitro-N-nitrosoguanidine

MMC: Mitomycin C

4NQO: 4-Nitroquinoline-1-oxide

9AA: 9-Aminoacridine

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

No significant increases in the frequency of revertant colonies were recorded for any of the bacterial strains, with any dose of the test material, either with or without metabolic activation. The test material was considered to be non-mutagenic under the conditions of this test.