Calcium cyanamide

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:

1979

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Remarks:

Animals were analysed for micronuclei six hours after the last treatment. Only one dosage was tested.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1979

Report date:

1979

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

yes

Remarks:

Animals were killed 6 hours after the last treatment; Only one dosage tested

GLP compliance:

not specified

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

Calcium cyanamide

EC Number:

205-861-8

EC Name:

Calcium cyanamide

Cas Number:

156-62-7

Molecular formula:

CN2.Ca

IUPAC Name:

calcium cyanoazanediide

impurity 1

Reference substance name:

Calcium oxide

EC Number:

215-138-9

EC Name:

Calcium oxide

Cas Number:

1305-78-8

Molecular formula:

CaO

IUPAC Name:

oxocalcium

impurity 2

Reference substance name:

Carbon

EC Number:

231-153-3

EC Name:

Carbon

Cas Number:

7440-44-0

Molecular formula:

C

IUPAC Name:

carbon

impurity 3

Reference substance name:

Hematite (Fe2O3)

EC Number:

215-275-4

EC Name:

Hematite (Fe2O3)

Cas Number:

1317-60-8

Molecular formula:

Fe2O3

IUPAC Name:

diiron oxide

impurity 4

Reference substance name:

Urea

EC Number:

200-315-5

EC Name:

Urea

Cas Number:

57-13-6

Molecular formula:

CH4N2O

IUPAC Name:

urea

impurity 5

Reference substance name:

Silicon dioxide

EC Number:

231-545-4

EC Name:

Silicon dioxide

Cas Number:

7631-86-9

Molecular formula:

O2Si

IUPAC Name:

dioxosilane

impurity 6

Reference substance name:

Trisilicon tetranitride

EC Number:

234-796-8

EC Name:

Trisilicon tetranitride

Cas Number:

12033-89-5

Molecular formula:

N4Si3

IUPAC Name:

trisilicon tetranitride

impurity 7

Reference substance name:

Calcium dihydroxide

EC Number:

215-137-3

EC Name:

Calcium dihydroxide

Cas Number:

1305-62-0

Molecular formula:

CaH2O2

IUPAC Name:

calcium dihydroxide

impurity 8

Reference substance name:

Aluminium oxide

EC Number:

215-691-6

EC Name:

Aluminium oxide

Cas Number:

1344-28-1

Molecular formula:

Al2O3

IUPAC Name:

aluminium oxide

impurity 9

Reference substance name:

Cyanoguanidine

EC Number:

207-312-8

EC Name:

Cyanoguanidine

Cas Number:

461-58-5

Molecular formula:

C2H4N4

IUPAC Name:

2-cyanoguanidine

impurity 10

Reference substance name:

Calcium acetylide

EC Number:

200-848-3

EC Name:

Calcium acetylide

Cas Number:

75-20-7

Molecular formula:

C2Ca

IUPAC Name:

calcium ethynediide

impurity 11

Reference substance name:

unknown

IUPAC Name:

unknown

Test material form:

solid: particulate/powder

Details on test material:

Kalkstickstoff (calcium cyanamide, technical grade)

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Central Institute for the Breeding of Laboratory Animals TNO, Zeist, The Netherlands
- Age at study initiation: 4-5 weeks
- Weight at study initiation: mean bodyweight for animals treated as Kalkstickstoff was 98 g for males and 82 g for females (animals of the water control: 99 g for males and 79 g for females)
- Assigned to test groups randomly: animals were assigned to groups according to bodyweight
- Fasting period before study: 14-15 hours
- Housing: screen-bottomed cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: yes; the duration of this period is not stated in the report

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 24°C ± 1°C
- Humidity (%): 40-60 %
- Photoperiod (hrs dark / hrs light): 12 hours light / 12 hours dark

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

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

Details on exposure:

DIET PREPARATION
Aqueous solutions / suspensions of each of the test samples were administered twice with an interval of 24 hours by stomach tube at dose levels of 1/5 of the LD50 in 5 mL of water per kg body weight.

Duration of treatment / exposure:

Animals were exposed twice at 153 mg/kg bw of test substance with an interval of 24 hours. The dose level used was based on the LD50 for rats (765 mg/kg bw).

Frequency of treatment:

Twice; with an interval of 24 hours

Post exposure period:

Animals were killed by decapitation 6 hours after the last treatment.

Doses / concentrations

Remarks:

Doses / Concentrations:
153 mg/kg bw
Basis:
actual ingested

No. of animals per sex per dose:

Only one dosage applied (corresponding to 1/5 of the LD50 value); 5 animals per sex

Control animals:

yes, concurrent vehicle

Positive control(s):

Trenimon
- Justification for choice of positive control(s): Trenimon is a well-known mutagen in rats
- Route of administration: intraperitoneal injection
- Doses / concentrations: 0.0625 mg/kg bw in physiological saline; administered twice with an interval of 24 hours

Examinations

Tissues and cell types examined:

A total of 400 erythrocytes per slide were examined for the incidence of micronucleated erythrocytes and the ratio of poly- and normochromatic erythrocytes.

Details of tissue and slide preparation:

The bone marrow of the femora was flushed into centrifuge tubes containing fetal calf serum and centrifuged. The cells were then resuspended by gentle mixing with a pasteur pipette. A drop of the solution was placed on a slide cleaned with methanol overnight and spread with a haemocytometer cover glass. Five slides were prepared for each animal. The smears were air-dried, fixed in methanol and stained according to May-Grünwald Giemsa.

Evaluation criteria:

No data

Statistics:

No data

Results and discussion

Test results

Key result

Sex:

male/female

Genotoxicity:

negative

Toxicity:

no effects

Vehicle controls validity:

valid

Negative controls validity:

not specified

Positive controls validity:

valid

Additional information on results:

No mortality or abnormalities of condition or behaviour, attributable to treatment, were observed in any of the animals during the exposure period.
The incidence of micronucleated erythrocytes and the percentage of polychromatic erythrocytes in the controls (water) and the test group (calcium cyanamide) were comparable.

Any other information on results incl. tables

Mean numbers of micronucleated erythrocytes and percentage polychromatic erythrocytes in bone marrow of rats after treatment with "Kalkstickstoff"

Type of treatment	sex	Mean incidence of micronucleated cells per 2000 erythrocytes per rat (range)	Percentage of polychromatic erythrocytes ± SD
Vehicle control	male	4.4 (2–7)	74.2 ± 3.5
Kalkstickstoff		5.2 (4–7)	70.2 ± 7.5
Vehicle control	female	5.0 (2–8)	63.8 ± 5.1
Kalkstickstoff		4.8 (2–8)	61.9 ± 2.0
Positive control:
Solvent	male	3.8 (2–8)	69
Trenimon		22.6 (20–26)	42
Solvent	female	3.6 (3–5)	70
Trenimon		34.4 (27–38)	46

Applicant's summary and conclusion

Conclusions:

Interpretation of results: negative
The results of the in vivo micronucleus test did not reveal any evidence for mutagenic activity of Kalkstickstoff in male or female rats.

Executive summary:

A study was conducted in order to evaluate the mutagenic potential of Calcium Cyanamide technical grade (Kalkstickstoff) in the in vivo micronucleus assay.

Therefore, rats were treated twice at 153 mg/kg bw with an interval of 24 hours and were examined for the incidence of micronucleated erythrocytes and the ratio of poly- and normochromatic erythrocytes six hours after the last treatment.

No mortality or abnormalities of condition or behaviour, attributable to treatment, were observed in any of the animals during the exposure period. The incidence of micronucleated erythrocytes and the percentage of polychromatic erythrocytes in the control and treated group were comparable. In contrast, the positive control Trenimon induced an increase in micronucleated erythrocytes and a decrease in the percentage of polychromatic erythrocytes. Thus, no evidence for mutagenic activity was observed for Kalkstickstoff and it was therefore concluded that Kalkstickstoff was negative in the in vivo micronucleus assay under the conditions of the study.