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Toxicological information

Carcinogenicity

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Description of key information

Oral: discriminating dose = approximately 5 mg/kg bw /day, male/female, mice, pre-guideline study, Leonard 1977

Key value for chemical safety assessment

Carcinogenicity: via oral route

Link to relevant study records
Reference
Endpoint:
carcinogenicity: oral
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Non-GLP, available as unpublished report, minor restrictions in design and/or reporting but otherwise adequate for assessment.
Qualifier:
no guideline followed
Principles of method if other than guideline:
25 mice per sex/dose were daily exposed to 0, 5, and 20 ppm test substance via diet for a duration of 78 weeks. The tumour incidence was investigated.
GLP compliance:
no
Species:
mouse
Strain:
not specified
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Mean body weight measured in week 1 of the study: Control group, 27.16 g males and 21.40 g females; 5 ppm dose group, 23.60 g males and 21.96 g females; 20 ppm dose group, 27.84 g males and 24.96 g females
- Age at study initiation: Approx. 2 months
- Housing: 6 or 7 per cage, wire mesh cages (26 cm x 26 cm x 9.5)
- Diet: Pelleted powder 'O' containing: 26.34 % barley, 9.10 % maize meal, 18.30 % sussex ground oats, 18.08 % bran, 4.46 % white fish meal, 1.34 % yeast, 13.17 % dried skimmed milk powder, 8.71 % meat and bone meal, 0.45 % salt, and 0.04 % 'nuclo' vitamine mixture (A, D3 and B2), ad libitum.
- Water: ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): approximately 50%
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: feed
Vehicle:
other: Maize oil and malt extract
Details on exposure:
DIET PREPARATION
Diets were prepared in a 'Hobart' mixer using powder 'O' diet. The appropriate concentration of the test substance was added to the dry powder 'O' and mixed for ten minutes prior to the addition of the following ingredients: 4 kg powder 'O', 900 g malt extract, 100 mL maize oil. Water was added as required to obtain the correct consistency for pelleting by mincing machine, and the diet was then dried under electric wall heaters.

Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
78 weeks
Frequency of treatment:
daily
Remarks:
Doses / Concentrations:
0, 5 and 20 ppm
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
0, 1.25, and 5 mg/kg/day
Basis:
other: estimated intake
No. of animals per sex per dose:
25
Control animals:
yes, plain diet
Details on study design:
An additional five male and five female mice were dosed in parallel with each group for blood level estimations after one month duration. However only two male and two female animals were used from dose group 5 and 20 ppm. The remaining animals continued on the trial and the survivors were killed without necropsy (apart from 1 animal) at termination.
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes

BODY WEIGHT: Yes
Time schedule for examinations: weekly

FOOD CONSUMPTION AND COMPOUND INTAKE:
Food consumption was measured during the first week of dosing on all animals for seven days.

CLINICAL CHEMISTRY: Yes
Two male and two female mice from the 5 and 20 ppm dose groups which had medicated diet in parallel with the main test animals, were bled 45 days after dosing commenced. Serum concentrations of the test substance were determined.
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
With the exception of four mice from the control group, 4 mice from the 5 ppm dose group and two mice from the 20 ppm dose group, all animals were necropsied after euthanasia with coal gas.

HISTOPATHOLOGY: Yes
After macroscopic examination, samples of tissues were taken for microscopic examination. The following organs were placed in Zenker's 1994 fixative and stained with haematoxylin and eosin: Accessory sex organs (left epididymis, seminal vesicle and uterus), adrenal (L), bladder (urinary), gonad (left ovary or testis), heart, intestine (large and small), kidney (L), liver, lung, lymph node (abdominal), mammary gland, muscle, pancreas, spleen, stomach, thyroid, thymus, and any other abnormal tissues.
Statistics:
Body weight: Student's t-test
Details on results:
CLINICAL SIGNS AND MORTALITY
The animals were generally in good body condition. The male mice had a tendency to fight and due to lacerations, occasionally this resulted in the formation of abscesses. Several mice suffered from minor respiratory problems. One mouse from the 5 ppm dose group was destroyed with middle ear disease. After ten months, two mice from the 20 ppm dose group convulsed on the same day whilst being weighed. This occurred again six weeks later with one of the animals. Other observations reported were thought to be associated with the development of tumours. See any other information on results incl. tables for the mortality rate.

BODY WEIGHT AND WEIGHT GAIN
There was a statistically significant lowered body weight in both sexes at termination of the test and this appeared to be dose related. See any other information on results incl. tables for the statistical analysis of body weight.

FOOD CONSUMPTION AND COMPOUND INTAKE
The mean daily food consumption per mouse was as follows: Control group: males 5.3 g and females 3.9 g; 5 ppm dose group: males 4.9 g and females 3.7 g; 20 ppm dose group: males 4.9 g and females 4.5 g.

CLINICAL CHEMISTRY
Serum concentration of the test substance were determined for two males and two females in both dose groups. 5 ppm resulted in serum concentrations of 0.0144 and 0.0117 µg/mL in males and 0.0055 and 0.0096 µg/mL in females. 20 ppm resulted in serum concentrations of 0.0274 and 0.0110 µg/mL in males and 0.0089 and 0.0082 µg/mL in females.

PATHOLOGY
There were no consistent microscopic and macroscopic findings observed in the treated or control groups. See any other information on results incl. tables for the incidence of tumours.
Relevance of carcinogenic effects / potential:
No biologically significant difference between the control and treated mice.
Dose descriptor:
NOAEC
Effect level:
> 5 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: The total incidence of tumours in the highest dose group and incidence in particular organ systems showed no biologically significant difference between the control and treated mice.

Mortality rate

 

Time of death (months)

group

sex

0 - 6

7 - 12

13 - 18

Survived to end

Control diet

Male

1

2 (2)

4 (2)

18

 

Female

0

0

5

20

5 ppm

Male

0

2 (2)

7 (1)

16

 

Female

0

1

3 (1)

21

20 ppm

Male

2

1

5

17

 

Female

1 (1)

2

5 (1)

17

( ) = no necropsy

Mortality was slightly higher in the high dose group.

Statistical analysis of body weight (g)

Sex

Group I

Group II

Group III

No.

Mean

No.

Mean

P

No.

Mean

P

Male

18

58.4

16

55.8

NS

17

47.6

<0.001

Female

20

51.9

21

46.8

NS

17

42.8

<0.01

P = Probability (students ‘t’ test) compared with control mean.

NS = Not significant.

Incidence of tumours

Organ

males

 

 

control

5 ppm

20 ppm

control

5 ppm

20 ppm

No. examined

21/25

22/25

25/25

25/25

24/25

23/25

Bladder

0

0

0

0

0

1

Bone marrow

1

2

1

1

0

1

Liver

6

7

5

2

2

2

Lung

0

0

2

2

2

0

Lymph node

0

1

0

0

0

0

Mammary gland

0

0

0

0

1

0

Pancreas

0

0

0

0

1

0

Spleen

0

0

3

0

3

1

Testes

1

0

0

0

0

0

Thymus

1

0

0

1

1

1

Uterus

0

0

0

1

0

0

Vagina

0

0

0

1

0

0

Total

9

10

11

8

10

6

Conclusions:
The total incidence of tumours in the highest dose group and incidence in particular organ systems showed no biologically significant difference between the control and treated mice.
Executive summary:

25 mice per sex/dose were daily exposed to 0, 5, and 20 ppm (approximately 1.25 and 5 mg/kg bw/day) test substance via diet for a duration of 78 weeks. Food consumption was measured during the first week of dosing for seven days. The mean daily food consumption per mouse was as follows: control group: males 5.3 g and females 3.9 g, 5 ppm dose group: males 4.9 g and females 3.7 g, 20 ppm dose group: males 4.9 g and females 4.5 g. Serum concentration of the test substance were determined for two males and two females in both dose groups. 5 ppm resulted in serum concentrations of 0.0144 and 0.0117 µg/mL in males and 0.0055 and 0.0096 µg/mL in females. 20 ppm resulted in serum concentrations of 0.0274 and 0.0110 µg/mL in males and 0.0089 and 0.0082 µg/mL in females. There was a statistically significant lowered body weight in both sexes at termination of the test and this appeared to be dose related. The total incidence of tumours in the highest dose group and incidence in particular organ systems showed no biologically significant difference between the control and treated mice.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
5 mg/kg bw/day
Study duration:
subchronic
Species:
mouse
Quality of whole database:
non-GLP compliant pre-guideline study, klimisch 2

Carcinogenicity: via inhalation route

Endpoint conclusion
Endpoint conclusion:
no study available

Carcinogenicity: via dermal route

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

25 mice per sex/dose were daily exposed to 0, 5, and 20 ppm (approximately 1.25 and 5 mg/kg bw/day) test substance via diet for a duration of 78 weeks. Food consumption was measured during the first week of dosing for seven days. The mean daily food consumption per mouse was as follows: control group: males 5.3 g and females 3.9 g, 5 ppm dose group: males 4.9 g and females 3.7 g, 20 ppm dose group: males 4.9 g and females 4.5 g. Serum concentration of the test substance were determined for two males and two females in both dose groups. 5 ppm resulted in serum concentrations of 0.0144 and 0.0117 µg/mL in males and 0.0055 and 0.0096 µg/mL in females. 20 ppm resulted in serum concentrations of 0.0274 and 0.0110 µg/mL in males and 0.0089 and 0.0082 µg/mL in females. There was a statistically significant lowered body weight in both sexes at termination of the test and this appeared to be dose related. The total incidence of tumours in the highest dose group and incidence in particular organ systems showed no biologically significant difference between the control and treated mice (Leonard,1977).

Justification for classification or non-classification

Based on the results of a 78 week mice carcinogenicity study the test substance is not classified according to the EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008.