Formamide

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study with acceptable restrictions.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

yes

Remarks:

, peripheral blood was obtained from mice of the 90-day repeated dose study

GLP compliance:

yes

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

Formamide

EC Number:

200-842-0

EC Name:

Formamide

Cas Number:

75-12-7

Molecular formula:

CH3NO

IUPAC Name:

formamide

Details on test material:

- Name of test material (as cited in study report): Formamide
- Analytical purity: approx. 100%
The test material was examined using IR-, UV-/visible light spectroscopy, proton nuclear magnetic resonance spectroscopy, TC, GC, and HPLC.

Test animals

Species:

mouse

Strain:

B6C3F1

Sex:

male/female

Details on test animals or test system and environmental conditions:

cf. repeated oral dose study; section 7.5.1

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Vehicle /solvent used: water

Details on exposure:

- oral gavage study

Duration of treatment / exposure:

90 days

Frequency of treatment:

5 treatments/week

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 10, 20, 40, 80, and 160 mg/kg bw/d
Basis:
actual ingested

No. of animals per sex per dose:

10 per dose and sex

Control animals:

yes, concurrent vehicle

Positive control(s):

none

Examinations

Tissues and cell types examined:

Preipheral blood; normochromatic and polychromatic erythrocytes

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION:
Dose selection for the 90-day oral gavage study was based on preceding 14-day studies.


TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields):
Treatment: 13 weeks, 5 days/week.
Sampling: from peripheral blood at the end of the 0-day study.

DETAILS OF SLIDE PREPARATION:
A detailed discussion of this assay is presented by MacGregor et al. (1990). At the end of the 3‑month toxicity study, peripheral blood samples were obtained from male and female B6C3F1 mice. Smears were immediately prepared and fixed in absolute methanol. The methanol‑fixed slides were stained with acridine orange and coded.


METHOD OF ANALYSIS:
Slides were scanned to determine the frequency of micronucleated cells in 2,000 normochromatic erythrocytes (NCEs) in each of 10 animals per treatment group. In addition, the percentage of polychromatic erythrocytes (PCEs) in a population of 1,000 erythrocytes was determined as a measure of bone marrow toxicity.

Evaluation criteria:

In the micronucleus test, an individual trial is considered positive if the trend test P value is less than or equal to 0.025 or if the P value for any single dosed group is less than or equal to 0.025 divided by the number of dosed groups. A final call of positive for micronucleus induction is preferably based on reproducibly positive trials (as noted above). Ultimately, the final call is determined by the scientific staff after considering the results of statistical analyses, the reproducibility of any effects observed, and the magnitudes of those effects.

Statistics:

The results were tabulated as the mean of the pooled results from all animals within a treatment group plus or minus the standard error of the mean. The frequency of micronucleated cells among NCEs was analyzed by a statistical software package that tested for increasing trend over dose groups with a one‑tailed Cochran‑Armitage trend test, followed by pairwise comparisons between each dosed group and the vehicle control group (ILS, 1990). In the presence of excess binomial variation, as detected by a binomial dispersion test, the binomial variance of the Cochran‑Armitage test was adjusted upward in proportion to the excess variation.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Remarks:

no induction of micronuclei in peripheral blood erythrocytes

Toxicity:

yes

Remarks:

reduced mean terminal body weight in males (80 and 160 mg/kg bw) and females (40, 80, 160 mg/kg bw)

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

not applicable

Any other information on results incl. tables

Results of the Micronucleus test, performed after 65 oral gavage doses in the 90-day repeated dose test are tabulated below. Harvesting time was 24 h after the last dose (cf. attached document: Annex E, table E3)

Dose (mg/kg/d)	MN-NCE/1000 NCE (mean±SEM)	Trend Test (P-value)(&)	% PCE
Males
0	1.15±0.21		2.18±0.08
10	1.20±0.17	0.442	2.38±0.14
20	1.30±0.23	0.334	2.21±0.11
40	1.00±0.17	0.676	1.94±0.06
80	0.95±0.09	0.732	1.84±0.07
160	0.95±0.15	0.732	2.15±0.23
		p=0.849 (§)
Females
0	0.85±0.13		2.09±0.08
10	0.80±0.11	0.569	2.11±0.08
20	0.80±0.13	0.569	1.87±0.14
40	0.95±0.12	0.369	1.84±0.12
80	1.05±0.12	0.258	1.70±0.08
160	0.80±0.15	0.569	1.78±0.11
		p=0.461

MN = micronucleated erythrocyte

NCE = normochromatic erythrocyte

PCE = polychromatic erythrocyte.

(&) = Pairwise comparison with the vehicle controls, significant at P=0.005 (ILS, 1990)

(§) = Significance of micronucleated NCEs/1,000 NCEs tested by the one-tailed trend test, significant at P=0.025 (ILS, 1990)

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative

Executive summary:

Conclusion:

Formamide was negative in the mouse peripheral blood micronucleus assay when blood cells from male and female mice (10/dose/sex) of the 13-week repeated dose toxicity tests were examined at termination of the subchronic oral gavage study (NTP, 2008). The method is valid and comparable to OECD TG 474. The dose levels used were 0, 10, 20, 40, 80, and 160 mg/kg bw/day. The incidence of polychromatic erythrocytes was not significantly changed in males or females at any dose level, indicating the absence of formamide-induced bone marrow toxicity (NTP, 2008).