Methyl myristate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

09 Apr - 21 May 2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Report date:

2010

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Version / remarks:

adopted in 1997

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.10 (Mutagenicity - In Vitro Mammalian Chromosome Aberration Test)

Version / remarks:

adopted in 2008

Deviations:

no

GLP compliance:

yes

Type of assay:

in vitro mammalian chromosome aberration test

Test material

Test material information

Constituent 1

Reference substance name:

Methyl laurate

EC Number:

203-911-3

EC Name:

Methyl laurate

Cas Number:

111-82-0

Molecular formula:

C13H26O2

IUPAC Name:

methyl laurate

Method

Target gene:

not applicable

Species / strain

Species / strain / cell type:

lymphocytes: cultured peripheral human lymphocytes

Details on mammalian cell type (if applicable):

- Type and identity of media: RPMI 1640 medium (Invitrogen Corporation)
- Other: blood was collected from healthy adult, non-smoking, male volunteers

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:

3h treatment: 33, 100 and 200 µg/mL
24h treatment: 100, 120 and 140 µg/mL
48h treatment: 30, 120 and 140 µg/mL

Vehicle / solvent:

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

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

ethanol

True negative controls:

no

Positive controls:

yes

Positive control substance:

cyclophosphamide

Remarks:

with S9-mix

Migrated to IUCLID6: 10 µg/mL

Controls 2

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

ethanol

True negative controls:

no

Positive controls:

yes

Positive control substance:

mitomycin C

Remarks:

without S9-mix

Migrated to IUCLID6: 0.5 µg/mL for a 3 h exposure period, 0.2 µg/mL for a 24 h exposure period and 0.1 µg/mL for a 48 h exposure period

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium

DURATION
- Exposure duration: 3, 24 and 48 h
- Fixation time (start of exposure up to fixation or harvest of cells): 3 h treatment: 24 and 48 h; 24h treatment: 24 h; 48 h treatment: 48 h

SPINDLE INHIBITOR (cytogenetic assays): colchicine 0.5 µL/mL medium
STAIN (for cytogenetic assays): Giemsa 5% (v/v) in tap water

NUMBER OF REPLICATIONS: 2

NUMBER OF CELLS EVALUATED: 100 per culture

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index of 1000 cells

OTHER EXAMINATIONS:
- Determination of polyploidy: yes

Evaluation criteria:

A test substance is considered positive (clastogenic) in the chromosome aberration test if: a) it induced a dose-related statistically significant (Chi-square test, one-sided, p < 0.05) increase in the number of cells with chromosome aberrations b) and a statistically significant and biologically relevant increase in the frequencies of the number of cells with chromosome aberrations was observed in the absence of a clear dose-response relationship.
A test substance is considered negative (not clastogenic) in the chromosome aberration test if none of the tested concentrations induced a statistically significant (Chi-square test, one-sided, p < 0.05) increase in the number of cells with chromosome aberrations. The preceding criteria are not absolute and other modifying factors might enter into the final evaluation decision.

Statistics:

Chi-square test, one-sided, p < 0.05

Results and discussion

Test results

Species / strain:

lymphocytes: cultured human peripheral lymphocytes

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: At a concentration of 200 µg/mL methyl laurate precipitated in the culture medium.

RANGE-FINDING/SCREENING STUDIES
The highest concentration analysed was selected based on the solubility of the test substance in the culture medium (3 h and 24 h continuous exposure) or on toxicity, i.e., inhibition of the mitotic index of about 50% or greater (48 h continuous exposure).

COMPARISON WITH HISTORICAL CONTROL DATA
The number of cells with chromosome aberrations found in the solvent control cultures was within the laboratory historical control data range.

Remarks on result:

other: strain/cell type: as specified above

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Table 1: Cytotoxic and genotoxic observations

Test item	Concentration	Mitotic Index	Aberrant cells in % (200 cells were scored)
	in µg/mL	in %	with gaps	without gaps
Exposure period 3h, fixation time 24h, without S9 mix
Ethanol	1.0% (v/v)	100	0.5	0.5
MMC	0.5	64	29*	29*
Test substance	33	90	0	0
	100	75	2	1.5
	200	66	2	1.5
Exposure period 3h, fixation time 24h, with S9 mix
Ethanol	1.0% (v/v)	100	0.5	0.5
CP	10	54	21*	21*
Test substance	33	100	0.5	0
	100	89	1	1
	200	66	0.5	0
Exposure period 24h, fixation time 24h, without S9 mix
Ethanol	1.0% (v/v)	100	2	1.5
MMC	0.2	54	47.3*^c)	46*^c)
Test substance	100	99	0	0
	120	80	2.5	2
	140	42	2.5	2
Exposure period 48h, fixation time 48h, without S9 mix
Ethanol	1.0% (v/v)	100	1	0.5
MMC	0.1	102	60*^b)	59*^b)
Test substance	30	93	0	0
	120	59	0.5	0.5
	140	44	1	0.5
Exposure period 3h, fixation time 48h, with S9 mix
Ethanol	1.0% (v/v)	100	1	1
CP	10	-^b)	52*^b)	51*^b)
Test substance	30	103	1	0.5
	100	101	1	0.5
	200	106	0	0

MMC: Mitomycin C

CP: Cyclophosphamide

^a): CP was fixed after 24 hours. The mitotic index could not be calculated as percentage of control

^b): 100 cells were scored

^c): 150 cells were scored

*: Significantly different from control group (Chi-square test), P < 0.001

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative