Fatty acids, tall-oil, reaction products with ethanolamine, ethoxylated

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2009-11-10 to 2010-04-26

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study according to OECD/EU guideline.

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 474 (Mammalian Erythrocyte Micronucleus Test)

Version / remarks:

July 21, 1997

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: Commission Regulation (EC) 440/2008, May 05,2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

61791-19-3

Cas Number:

61791-19-3

IUPAC Name:

61791-19-3

Test material form:

not specified

Test animals

Species:

mouse

Strain:

NMRI

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Hartan Laboratories B.V. Postbus 6174, 5960 AD Horst/ The Netherlands
- Age at study initiation: 8-12 weeks
- Weight at study initiation: mean value 39.3 g
- Assigned to test groups randomly: yes
- Fasting period before study: no
- Housing: single
- Diet (ad libitum): pelleted standard diet
- Water (ad libitum): tap water
- Acclimation period: minimum of 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 2
- Humidity (%): 28-65
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Vehicle used: corn oil
- Justification for choice of vehicle: relative non-toxicity for the animals
- Amount of vehicle: 10 mL/kg bw

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: On the day of the experiment, the test item was formulated in corn oil.

Duration of treatment / exposure:

The animals received the test item, the vehicle or the positive control substance once orally.

Frequency of treatment:

The animals received the test item, the vehicle or the positive control substance once orally.

Post exposure period:

24 and 48 hours

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
2000, 1000 and 500 mg/kg bw
Basis:
actual ingested
24 hour preparation interval

Doses / concentrations 2

Remarks:

Doses / Concentrations:
2000 mg/kg bw
Basis:
actual ingested
48 hour preparation interval

No. of animals per sex per dose:

7 male mice/dose/sampling time

Control animals:

yes, concurrent vehicle

Positive control(s):

cyclophosphamide
- Route of administration: orally, once
- Dose: 40 mg/kg bw

Examinations

Tissues and cell types examined:

erythrocytes from bone marrow

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION: The maximum tolerated dose level is determined to be the dose that causes toxic reactions without having major effects on survival within 48 hours.

TREATMENT AND SAMPLING TIMES: Sampling of the bone marrow was done 24 and 48 hours after treatment, respectively.

DETAILS OF SLIDE PREPARATION:
The animals were sacrificed using CO2 followed by bleeding. The femora were removed, the epiphyses were cut off and the marrow was flushed out with foetal calf serum using a syringe. The cell suspension was centrifuged at 1500 rpm (390 x g) for 10 minutes and the supernatant was discarded. A small drop of the re-suspended cell pellet was spread on a slide. The smear was air-dried and then stained with May-Grünwald/Giemsa. Cover slips were mounted with EUKITT. At least on slide was made from each bone marrow sample.

METHOD OF ANALYSIS:
Evaluation of the slides was performed using NIKON microscopes with 100 x oil immersion objectives. Per animal 2000 polychromatic erythrocytes (PCE) were analysed for micronuclei. To describe a cytotoxic effect the ratio between polychromatic and normochromatic erythrocytes was detetmined in the same sample and expressed in polychromatic erythrocytes per 2000 erythrocytes. The analysis was performed with coded slides.
All animals per test group were evaluated as described.

Evaluation criteria:

A test item is classified as mutagenic if it induces either a dose-related increase or a clear increase in the number of micronucleated polychromatic erythrocytes in a single dose group.
A test item that fails to produce a biological relevant increase in the number of micronucleated polychromatic erythrocytes is considered as non-mutagenic in this system.

Statistics:

Nonparametric Mann-Whitney test

Results and discussion

Test results

Sex:

male

Genotoxicity:

negative

Toxicity:

yes

Remarks:

ruffled fur

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

RESULTS OF RANGE-FINDING STUDY
- Dose: 2000 mg/kg bw
- Solubility: in corn oil
- Clinical signs of toxicity in test animals: ruffled fur
- Evidence of cytotoxicity in tissue analyzed: no
- Harvest times: 1 h, 2-4 h, 6 h, 24 h, 30 h, and 48 h after administration

RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei: No
- Statistical evaluation: Negative control group versus test group
500 mg/kg bw (24h): p 0.3674, not significant
1000 mg/kg bw (24h): p 0.1777, not significant
2000 mg/kg bw (24h): p 0.2716, not significant
2000 mg/kg bw (48h): p 0.3817, not significant
Positive control (24h): 0.0003, significant

Any other information on results incl. tables

Summary of Micronucleus Test Results

test group	dose (mg/kg bw)	sampling time (h)	PCEs with micronuclei (%)	range	PCE per 2000 erythrocytes
vehicle	0	24	0.064	1 - 2	1134
test item	500	24	0.079	0 - 3	1107
test item	1000	24	0.093	0 - 3	1123
test item	2000	24	0.086	0 - 3	1134
positive control	40	24	2.286	21 - 73	1101
test item	2000	48	0.079	0 - 3	1177