Amines, C16-18 (even numbered)-alkyl , salts with phosphoric acid, mono- and di-C16-18 (even numbered) alkyl esters

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / bone marrow chromosome aberration

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

1989

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1989

Report date:

1989

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 475 (Mammalian Bone Marrow Chromosome Aberration Test)

Principles of method if other than guideline:

Study was performed before actual guideline was adopted (21 Jul 1997).

GLP compliance:

yes

Type of assay:

chromosome aberration assay

Test material

Test material information

Constituent 1

Reference substance name:

(Z)-octadec-9-enylamine

EC Number:

204-015-5

EC Name:

(Z)-octadec-9-enylamine

Cas Number:

112-90-3

Molecular formula:

CH3(CH2)7CH=CH(CH2)7CH2NH2

IUPAC Name:

octadec-9-en-1-amine

Constituent 2

Reference substance name:

(Z)-octadec-9-en-1-amine

IUPAC Name:

(Z)-octadec-9-en-1-amine

Constituent 3

Reference substance name:

Oleyl Alkylamines

IUPAC Name:

Oleyl Alkylamines

Details on test material:

- Name of test material (as cited in study report): Oleylamine

Test animals

Species:

mouse

Strain:

ICR

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan Sprague Dawley, Inc., Frederick , MD, USA
- Age at study initiation: 6-8 weeks old
- Weight at study initiation: males: 28-38 g; females: 22-28 g
- Assigned to test groups randomly: The animals were assigned to 13 experimental groups of 5 males and 5 females each based on a computer-generated random number table.
- Housing: The animals were housed in plastic autoclavable cages with filter tops during quarantine and the experimental period. Hardwood chips were used for bedding.
- Diet (e.g. ad libitum): certified laboratory rodent chow which had been analysed for environmental contaminants; ad libitum
- Water (e.g. ad libitum): tap water; ad libitium

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-25 °C
- Humidity (%): 50±20
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Vehicle(s)/solvent(s) used: corn oil
- Amount of vehicle (if gavage or dermal): 10 mL/kg bw
- Lot/batch no. (if required): Sept1389J (Giant Foods, Washington, D.C., USA)

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
The test article was dissolved in a total volume of 10 mL/kg bw.

Duration of treatment / exposure:

single administration

Frequency of treatment:

single administration

Post exposure period:

6, 12, or 24 h

Doses / concentrations

Remarks:

Doses / Concentrations:
500, 2500, 5000 mg/kg
Basis:
nominal conc.

No. of animals per sex per dose:

5

Control animals:

yes, concurrent vehicle

Positive control(s):

triethylenemelamine: dissolved in sterile distilled water at a concentration of 0.05 mg/mL
- Route of administration: injection

Examinations

Tissues and cell types examined:

bone marrow cells

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION:
Based on the preliminary dose-finding study, the doses for the main study were selected.

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields):
Colchicine, used to arrest dividing cells at metaphase, was administered ip at 1 mg/kg to all mice two to four hours prior to sacrifice. Animals were sacrificed by carbon dioxide asphyxiation. Immediately following sacrifice, the femur was exposed and the bone marrow was aspirated.

DETAILS OF SLIDE PREPARATION:
The cells were centrifuged and resuspended in a fixative (methanol:acetic acid, 3:1, v/v). The cell suspension was dropped onto a wet glass slide and dried. 3 slides were prepared from each animal. The dry slides were stained with 5% Giemsa stain.

METHOD OF ANALYSIS:
A minimum of 50 metaphase cells containing 40±2 centromeres were examined from each animals and scored for chromatid-type and chromosome-type aberrations. Chromatid and isochromatid gaps were recorded but not included in the analysis. The mitotic index was recorded as the percentage number of cells in mitosis based upon 500 cells counted per animals.

Evaluation criteria:

The test article was considered to induce a positive response when the number of aberrant cells was significantly increased in a dose responsive manner relative to the vehicle control. A significant increase at the high dose only with no dose-response was considered suspect.
The percentage of cells in the solvent control group demonstrating aberrations of any type, other than gaps, must not exceed 4%. The cells with aberrations in the positive control group must be statistically increased (p=0.05, Fisher´s Exact Test).

Statistics:

Fisher´s Exact Test; Cochran-Armitage trend test

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

yes

Remarks:

One female receiving 2500 mg/kg died prior to colchicine administration.

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

RESULTS OF RANGE-FINDING STUDY
- Dose range: 763, 1221, 1953, 3125, 5000 mg in corn oil/kg bw
- Clinical signs of toxicity in test animals: diarrhea, ruffled fur, crusty eyes and hunched posture (one female receiving 5000 mg/kg, one female receiving 3125 mg/kg, one female receiving 1221 mg/kg were found dead)
- Other: A single administration of 5000 mg/kg was selected as the high dose for the cytogenetics study.

RESULTS OF DEFINITIVE STUDY
- The marginal increases in percentage of aberrant cells observed in the high dose group were not statistically significant (p>0.05, Fisher´s exact test). In fact, the percentage of damaged cells was not significantly increased in the Oleylamine-treated animals, regardless of sex, dose or sacrifice time (p>0.05, Fisher´s exact test).
One female receiving 2500 mg/kg died prior to colchicine administration. No significant reduction in the rate of body weight gain was observed. Clinical observations in treated animals included diarrhea, piloerection, lethargy, crusty eyes and irregular breathings.

Applicant's summary and conclusion

Conclusions:

Under the conditions of the assay, Oleylamine at dose levels of 500, 2500 and 5000 mg/kg body weight was negative in the in vivo cytogenetic assay using male and female ICR mice. The negative and positive controls fulfilled the requirements for determination of a valid test.

Executive summary:

A chromosomal aberration test in mice bone marrow cells with octadecenylamine ("Oleylamine") was performed. Groups of five male and five female mice were administered single doses of 500, 2500 or 5000 mg/kg bw in corn oil by oral gavage at a volume of 10 ml/kg bw (purity >90%). Bone marrow cells were collected 6, 12 and 24 hours after treatment. No increases in percentages of aberrant cells were observed in the test substance treated animals regardless of dose or harvest time. One female in the 2500 mg/kg bw group died prematurely. No significant reduction in the rate of body weight gain was observed. Clinical signs of toxicity were observed in test substance-treated mice indicating that the test substance was systemically available after oral application. Based on the test results, oleylamine was negative in this in vivo mutagenicity assay. The concurrent negative and positive controls fullfilled the requirements for a valid test.