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Repeated dose toxicity: oral

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Administrative data

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
November 8, 2004 - February 23, 2005
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
There are no deviations from the recommended guideline. Meets the requirements of GLP

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2005
Report date:
2005

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral))
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
A mixture of: propan-2-one-O,O'(methoxyvinylsilandiyl)dioxime; propan-2-one-O-(dimethoxyvinylsilyl)oxime; propan-2-one-O,O',O''-(vinylsilantriyl)trioxime
EC Number:
458-680-3
EC Name:
A mixture of: propan-2-one-O,O'(methoxyvinylsilandiyl)dioxime; propan-2-one-O-(dimethoxyvinylsilyl)oxime; propan-2-one-O,O',O''-(vinylsilantriyl)trioxime
Cas Number:
797751-44-1
Molecular formula:
not applicable, multiconstituent substance
IUPAC Name:
3-ethenyl-3-methoxy-6-methyl-2,4-dioxa-5-aza-3-silahept-5-ene; 5-ethenyl-2,8-dimethyl-5-{[(propan-2-ylidene)amino]oxy}-4,6-dioxa-3,7-diaza-5-silanona-2,7-diene; 5-ethenyl-5-methoxy-2,8-dimethyl-4,6-dioxa-3,7-diaza-5-silanona-2,7-diene
Test material form:
liquid

Test animals

Species:
rat
Strain:
other: Fischer, F344/NHsd, SPF.
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Harlan Winkelman
- Age at study initiation: about 7 weeks
- Weight at study initiation: males about 155 g; females about 110 g. All the animals were within ±20% of the mean weight per sex at the time of allocation groups.
- Housing: group caging, Makrolon cages type IV
- Diet (e.g. ad libitum): ad libitum. Exception: feed was withdrawn the day before blood sampling
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 7 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): average of 21.9 ºC
- Humidity (%): average of 45.4 ºC
- Air changes (per hr): 12 per hour
- Photoperiod (hrs dark / hrs light): artificial light from 6 a.m. to 6 p.m.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:
The test substance was blended with the vehicle (corn oil). Preparations of the test substance were made freshly every day shortly before the administration to the animals. Appropriate preparations were made to allow a uniform dose volume for all groups.

DIET PREPARATION
- Mixing appropriate amounts with (Type of food): Altromin 1324 forte, gamma irradiated with 25kGy60Co

VEHICLE
- Justification for use and choice of vehicle (if other than water): as the test substance is known to react immediately with water, corn oil was chosen as solvent.
- Amount of vehicle (if gavage): 5 mL per kg body weight
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Determinations of the test substance in some selected preparations for stability, concentration and homogeneity were performed by gas chromatography with flame ionisation detection (GC/FID).
Concentration test: A deviation of the mean of the 3 samples from the target concentration of at most ± 10 % was tolerated.
Homogeneity test: A deviation of the individual samples from the mean of at most ± 10 % was tolerated.
Duration of treatment / exposure:
Test duration: 28 days
Frequency of treatment:
Once a day for 28 consecutive days.
Doses / concentrationsopen allclose all
Dose / conc.:
20 mg/kg bw/day (actual dose received)
Dose / conc.:
63 mg/kg bw/day (actual dose received)
Dose / conc.:
200 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
Low dose (20 mg/kg/day): 10 animals (5 males and 5 females).
Mid dose (63 mg/kg/day): 10 animals (5 males and 5 females).
High dose (200 mg/kg/day): 10 animals (5 males and 5 females) and an additional group of 10 animals (5 males and 5 females) for observation of reversibility of substance effects.
Control animals:
yes, concurrent no treatment
yes, concurrent vehicle
Details on study design:
- Dose selection rationale:
the doses were derived from the results of the dose range findings study.
Groups of 5 male and 5 female rats were given doses of 100 mg or 316 mg or 1000 mg test substance per kg body weight for 7 consecutive days. Investigations: Body weights, feed consumption, animal observation, gross pathology.
Results: 1 high dosed female died early, with the death related to the test substance. Gross necropsy findings gave some indication for hepatotoxicity.Body weights of the mid and high dosed males and high dosed females were significantly reduced. Feed consumption was reduced in parallel.
Therefore 200 mg test substance per kg body weight, interpolated between low and mid dose of the dose range finding study, are chosen as high dose for the main study. 20 mg test substance per kg, i.e. 10 % of the high dose, is chosen as the low dose, where no toxic effects are anticipated. The middose is interpolated geometrically.

- Post-exposure recovery period in satellite groups: 14 days

Examinations

Observations and examinations performed and frequency:
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: all animals, once a week

BODY WEIGHT: Yes
- Time schedule for examinations: all animals, once a week

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: Yes

HAEMATOLOGY: Yes
- Time schedule for collection of blood: all the animals groups on day 29, except the dose satellite group and the control satellite group that the blood was collected 14 days after (on day 43).
- Anaesthetic used for blood collection: Yes (ether)
- Animals fasted: Yes
- How many animals: 30 animals
- Parameters checked in table [No.3] were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: all the animals groups on day 29, except the dose satellite group and the control satellite group that the blood was collected 14 days after (on day 43).
- Animals fasted: Yes
- How many animals: 30 animals
- Parameters checked in table [No.9] were examined.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

OTHER: organ weights (see table 4, summary data)
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
Statistics:
Analysis of variance followed by the Scheffé-test, t-test and H-test of Kruskal and Wallis followed by the test of Nemenyi were used

Results and discussion

Results of examinations

Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
One single animal (contral satellite graup) was found to suffer fram an eye lesion. This is not related to the test substance.
Mortality:
no mortality observed
Description (incidence):
No test substance related death occurred during the study. All animals survived until their scheduled sacrifice.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
High dosed satellite group males had a significantly lower body weight than the controls in the last week of dosing and in the first week of the recovery period. Feed consumption of the high dosed males was marginally impaired.
The body weight gain of the high dosed satellite group males, body weight gain was significantly reduced in the first week and the fourth week of dosing and significantly elevated in the first week of the recovery period.
There were no significant differences in the body weight gain of the females.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
In the males, the high dosed and the high dosed satellite group had a slightly lower feed consumption during the dosing period, followed by a slightly higher feed consumption at the end of the recovery period, both when compared with the controls. The differences were not markedly pronounced and are given only borderline relevance.
There were no noteworthy differences or dose related trends noted in the feed consumption of the females.
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not specified
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
The test substance causes damage to mature erythrocytes in the peripheral blood, resulting in alterations in virtually all erythrocyte-related parameters at the haematological examination. In parallel to this damage there is a marked repair by ongoing by an increased new production of young erythrocytes, notable in an increased proportion of polychromatic erythrocytes in the peripheral blood and also in haematopoiesis in the spleen, inducing even a significant weight increase of this organ, and - less pronounced - a hypercellularity of the bone marrow. The repair was efficient enough to allow the animals to live without physical behavioural abnormalities. There was no indication for a negative effect to the blood cell production in the bone marrow found.
Effects were similarly noted in males and females at the same doses, no sex difference in the response to the test substance can be derived from the results.
Effects were only partly reversible during the recovery period, though there was a clear tendency towards a return to normal.
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
Some haematological changes in neutrophil and lymphocyte counts and some clinical biochemical changes and the increased liver weights remain without a clear interpretation. The haematological changes may be secondary to the erythrocyte alterations, the biochemical ones may indicate a borderline effect to the liver.
Effects were similarly noted in males and females at the same doses, no sex difference in the response to the test substance can be derived from the results.
Effects were only partly reversible during the recovery period, though there was a clear tendency towards a return to normal.
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
All observations made are within anormal behavioural pattern of rats of the strain and age used. Reactivity, reflexes and grip strength were also adequate.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
The liver and the spleen weight differences were attributed to the test substance.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
In all high dosed animals a large spleen was noted. This alteration is attributed to the test substance.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Histopathologically, extramedullary haematopoiesis was found in the spleens of all high dosed animals and most mid dosed animals. Hypercellularity of the bone marrow may be also due to the test substance.
Histopathological findings: neoplastic:
no effects observed
Other effects:
no effects observed

Effect levels

Key result
Dose descriptor:
NOEL
Effect level:
20 mg/kg bw/day (nominal)
Based on:
test mat. (dissolved fraction)
Sex:
male/female
Basis for effect level:
haematology
histopathology: non-neoplastic

Target system / organ toxicity

Key result
Critical effects observed:
no

Any other information on results incl. tables

Mean body weight gain (Table 1)

Data are presented by mean, standard deviation (sd) and number of animals (n). Significant differences to the control group are indicated by an underline number.

Table1: Males

group

body weight gain (g/day per animal) for Days

 

-5-1

1-8

8-15

15-22

22-28 28-36

36-42

K mean

4.90

3.94

3.77

3.03

2.47

 

-

sd

0.62

0.54

0.22

0.31

0.76

 

 

n

5

5

5

5

5

 

 

KS mean

5.17

4.17

3.74

3.09

2.83

2.08

1.30

sd

0.58

0.59

0.47

0.59

0.47

0.52

0.68

n

5

5

5

5

5

5

5

A mean

4.47

4.06

3.29

2.77

1.90

-

-

sd

0.66

0.62

0.54

0.30

0.25

 

 

n

5

5

5

5

5

 

 

B mean

4.47

3.60

3.17

3.03

2.90

-

-

sd

0.59

0.31

0.65

0.59

0.38

 

 

n

5

5

5

5

5

 

 

C mean

4.97

3.49

3.03

2.49

1.93

-

-

sd

0.84

0.81

0.75

0.61

0.45

 

 

n

5

5

5

5

5

 

 

CS mean

4.67

3.31

3.37

2.74

1.70

2.83

1.97

sd

0.42

0.37

0.44

0.41

0.61

0.17

0.49

n

5

5

5

5

5

5

5

Table 1 cont.: Females

group

body weight gain (g/day per animal) for Days

 

 

-5-1

1-8

8-15

15-22

22-28

28-36

36-42

K

mean

2.77

1.71

1.63

1.34

0.78

-

-

 

sd

0.62

0.30

0.16

0.46

0.19

 

 

 

n

5

5

5

5

5

 

 

KS

mean

2.90

1.66

2.00

1.34

0.95

0.58

0.03

 

sd

0.56

0.48

0.29

0.26

0.53

0.17

0.52

 

n

5

5

5

5

5

5

5

A

mean

2.57

1.74

2.00

1.31

0.88

-

-

 

sd

0.58

0.31

0.27

0.48

0.54

 

 

 

n

5

5

5

5

5

 

 

B

mean

2.97

2.06

1.77

1.23

1.23

-

-

 

sd

0.25

0.33

0.19

0.33

0.55

 

 

 

n

5

5

5

5

5

 

 

C

mean

2.30

1.54

1.89

1.54

0.83

 

-

 

sd

0.40

0.51

0.46

0.49

0.46

 

 

 

n

5

5

5

5

5

 

 

CS

mean

2.37

1.57

2.00

1.20

0.10

1.78

0.50

 

sd

0.75

0.17

0.39

0.16

1.39

1.03

0.60

 

 

5

5

5

5

5

5

5

Clinical Biochemistry Data (Table 2)

parameter (sex)

low dose

mid dose

high dose

high dose
after recovery

albumin (males)

-

-

x

-

cholesterol (males)

-

-

x

-

total protein (males)

-

-

x

-

urea (males)

-

-

-

+

alanin aminotransferase (females)

-

-

x

-

alkaline phosphatase (females)

-

-

-

+

calcium (females)

-

-

-

x

glucose (females)

-

-

-

x

key to the symbols:
-: no significant difference to the control, +: significant increase, x: significant decrease.

All significant differences in clinical biochemistry are attributed to the test substance.

Haematology (Table 3)

parameter (sex)

low dose

mid dose

high dose

high dose
after recovery

red blood cell count (males)

-

X

X

X

red blood cell count (females)

-

X

X

X

haemoglobin concentration (males)

-

X

X

+

haemoglobin concentration (females)

-

X

X

-

haematocrit (males)

-

X

X

+

haematocrit (females)

-

-

X

-

mean corpuscular haemoglobin, pg (males)

-

-

+

+

mean corpuscular haemoglobin, pg (females)

-

+

+

+

mean corpuscular haemoglobin, g/dL (males)

-

-

X

-

mean corpuscular haemoglobin, g/dL (females)

-

-

X

-

mean cell volume (males)

-

-

+

+

mean cell volume (females)

-

+

+

+

white blood cell count (males)

-

-

+

-

white blood cell count (females)

-

-

+

-

monocytes, % (males)

-

-

-

+

eosinophils, % (males)

 

 

 

X

lymphocytes, n (males)

-

-

+

-

lymphocytes, n (females)

-

-

+

-

prothrombin time (males)

-

-

-

X

neutrophils, n (males)

-

-

+

-

neutrophils, n (females)

-

-

-

+

monocytes, n (males)

-

-

+

+

eosinophils, % (males)

-

-

-

X

platelet count (females)

-

-

+

-

polychromatic erythrocytes (%, males)

-

-

+

-

polychromatic erythrocytes (%, females)

-

-

+

-

key to the symbols:
-: no significant difference to the control, +: significant increase, x: significant decrease

 

All significant differences in haematology are attributed to the test substance.

Necropsy findings

In all high dosed animals a large spleen was noted. This alteration is attributed to the test substance.

Organ weights(Table 4)

Survey of the results:

parameter
(sex)

low dose

mid dose

high dose

high dose
after recovery

spleen (males, absolute weight)

-

-

+

-

spleen (females, absolute weight)

-

-

+

+

liver (females, absolute weight)

-

-

-

+

liver (males, organ weight/body weight ratio)

-

-

+

+

spleen (males, organ weight/body weight ratio)

-

-

+

+

liver (females, organ weight/body weight ratio)

-

-

-

+

spleen (females, organ weight/body weight ratio)

-

-

+

+

spleen (males, organ weight/brain weight ratio)

-

-

+

-

liver (females, organ weight/brain weight ratio)

-

-

-

+

spleen (females, organ weight/body weight ratio)

-

-

+

+

key to the symbols:
-: no significant difference to the control, +: significant increase, x: significant decrease

The liver and spleen weight differences are attributed to the test substance.

Histopathology

The presence of extramedullary haematopoiesis in the spleens of all high dosed and almost all mid dosed animals of both sexes is related to the test substance. A minor pronounced effect, similar to that mentioned before, is hypercellularity in the bone marrow, noted in a few animals without gaining statistical significance.

All other histopathological findings are not related to the test substance and considered as spontaneous alterations without toxicological relevance.


Applicant's summary and conclusion

Conclusions:
The No-observed-effect-level of the test substance was at 20 mg per kg body weight and day in both sexes.
Executive summary:

The Repeated dose 28-Day Oral toxicity Study in Rodents for the test substance was performed in F344 rats. The test material was administered blended with corn oil, given orally by gavage to 3 groups of 5 male and 5 female rats, once a day for 28 consecutive days. The dose used were 20, 63, and 200 mg/kg/day. A negative control group (treated with the vehicle) was included. In addition, 2 groups of 5 males and 5 females each (i.e. one high dose satellite group and one control satellite group) were treated in the same way as their corresponding groups, but were kept for further 14 days without test substance administration in an attempt to observe the reversibility or persistence of test substance induced lesions.

Investigations:

Clinical observations and body weights: all animals, once a week.

Functional observations: all animals, once, in the last week of the dosing period.

Haematology, clinical biochemistry,necropsy with gross pathological examinationand organ weight determination: all animals on Day 29 and the satellite group animals on day 43.

Histopathological examination: all animals of high dose group and control group, and the spleens of all animals.

The effects noted at a dose of 200 mg/kg and below were mainly adaptations to damage to peripheral erythrocytes without an impairment of the blood cell production in the bone marrow. All other effects noted were only borderline ones.

The No-observed-effect-level of the test substance was at 20 mg per kg body weight and day in both sexes.