Fatty acids, tall-oil, compds. with diethanolamine

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Type of information:

experimental study

Adequacy of study:

key study

Study period:

No data

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1985

Report date:

1985

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

not specified

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EEC Directive 79/831, Annex V, Method No. 431

Deviations:

not specified

Principles of method if other than guideline:

Not applicable

GLP compliance:

not specified

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

Amides, C8-18 (even numbered) and C18-unsatd., N,N-bis(hydroxyethyl)

EC Number:

931-329-6

Cas Number:

68155-07-7

Molecular formula:

The alkyl chain length of the amide ranges between 8 and 18 carbon atoms

IUPAC Name:

Amides, C8-18 (even numbered) and C18-unsatd., N,N-bis(hydroxyethyl)

Constituent 2

Reference substance name:

2,2'-iminodiethanol

EC Number:

203-868-0

EC Name:

2,2'-iminodiethanol

Cas Number:

111-42-2

Molecular formula:

C4H11NO2

IUPAC Name:

2,2’-iminodiethanol

Test material form:

liquid

Test animals

Species:

mouse

Strain:

NMRI

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Breeders were purchased from G1. Bomholtgard Ltd., but the mice used were born in the Scantox Laboratories
- Age at study initiation: 6-7 wk
- Weight at study initiation: 25-29 g
- Assigned to test groups randomly: Yes, in 5 groups
- Fasting period before study: No
- Housing: 5 animals/cage, males and females separately in type III Macrolone cages, bedding used was special softwood sawdust "Spanvall Special White " from Spanvall Ltd., DK-4535 Vallekilde
- Diet: Complete rodent diet "Altromin 1314" from Chr. Petersen Ltd., DK-4100 Ringsted, ad libitum
- Water (e.g. ad libitum): Drinking water adjusted to pH 2.5 with hydrochloric acid

ENVIRONMENTAL CONDITIONS
- Temperature: 21±2 °C
- Humidity (%): 55 ± 15%
- Air changes: 10 per hr
- Photoperiod: 12 hrs dark/12 hrs light

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Vehicle(s)/solvent(s) used: Distilled water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: Dilution of the test material in distilled water

Duration of treatment / exposure:

Not applicable

Frequency of treatment:

Once

Post exposure period:

24, 48 and 72 h

Doses / concentrations

Dose / conc.:

15 000 mg/kg bw/day (nominal)

Remarks:

equivalent to 14.7 mL/kg

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Positive control(s):

cyclophosphamide
- Route of administration: Oral
- Doses / concentrations: 30 mg/10 mL

Examinations

Tissues and cell types examined:

Bone marrow erythrocytes

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION: Preliminary investigations - A few mice were treated orally by various concentrations of the test material diluted with distilled water. Thereby the maximum tolerated dose was estimated at 15 g/kg bw. At this dosage bone marrow smears showed a reduced number of polychromatic erythrocytes (PCE) as compared with normochromatic erythrocytes (NCE). At a dose exceeding 15 g/kg bw the mortality was too high.

DETAILS OF SLIDE PREPARATION: Immediately after sacrifice, femurs of a mouse were dissected free of muscle, and by a 1 mL syringe with needle the bone marrow was flushed out into 5 mL of fetal calf serum. After thorough shaking, the mixture was centrifuged for 10 min. at about 1000 rpm. Thereafter, smears were made after removal of the supernatant. The specimens were fixed in methanol and stained with May-Grunwald/Giemsa.

METHOD OF ANALYSIS: Prior to microscopic assessment, all slides were furnished with code numbers, so that the counting was blind. The following counts were made:
Number of normochromatic erythrocytes (NCE) per 1000 erythrocytes
Number of polychromatic erythrocytes (PCE) per 1000 erythrocytes
Number of micronuclei (MN) in 1000 normochromatic erythrocytes
Number of micronuclei (MN) in 1000 polychromatic erythrocytes.

Evaluation criteria:

Increase in the frequency of micronucleated polychromated erythrocytes in treated animals as compared to controls.

Statistics:

The statistical difference was analysed by one-way ANOVA. In the case of PCE (%) the test was performed on the values observed, and for the MN (per thousand) the test was done on computed rank values transformed to normal scores according to Blom's method. (G. Blom Statistical Estimates and Transformed Beta Variables. New York: John Wiley and Sons, Inc., 1958).

Results and discussion

Test results

Key result

Sex:

male/female

Genotoxicity:

negative

Toxicity:

not specified

Vehicle controls validity:

valid

Negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

RESULTS OF RANGE-FINDING STUDY
- Dose range: Up to 15 g/kg bw
- Clinical signs of toxicity in test animals: Maximum tolerated dose was estimated at 15 g/kg bw. At a dose exceeding 15 g/kg bw, the mortality was too high.
- Evidence of cytotoxicity in tissue analyzed: At 15 g/kg bw, bone marrow smears showed a reduced number of polychromatic erythrocytes (PCE) as compared with normochromatic erythrocytes (NCE).

RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei (for Micronucleus assay): No significant difference as compared to controls.
- Appropriateness of dose levels and route: Yes
- Statistical evaluation: Yes

Any other information on results incl. tables

For detailed results and bar charts kindly refer to the attached background materials section of the IUCLID.

Applicant's summary and conclusion

Conclusions:

Under the study conditions, the test substance did not induce an increase in the frequency of micronuclei in bone marrow cells of the mouse, and was therefore considered to be non-mutagenic.

Executive summary:

A study was conducted to evaluate the in vivo genetic toxicity of the test substance, C8-18 and C18-unsatd. DEA, according to the method recommended in the First Addendum to OECD Guideline 474 and Method No. 431, Annex V of EEC Directive 79/831. The experimental animals were 70 NMRI mice, divided into 5 groups. Of the 5 groups, 3 were test groups, 1 negative control group and 1 positive control group. The test groups were treated with 15000 mg test substance/kg bw, the negative control group with distilled water and the positive control group with 30 mg cyclophosphamide/kg bw. The mice were killed 24, 48 and 72 h, respectively, after treatment. From bone marrow smears, micronucleus counts were made per 1000 polychromatic erythrocytes. The test substance did not induce an increase in the frequency of micronucleated normochromatic erythrocytes in peripheral blood samples from both male and female mice. Under the study conditions, the test substance did not induce an increase in the frequency of micronuclei in bone marrow cells of the mouse, and was therefore considered to be non-mutagenic (Kallesen, 1985).