Registration Dossier

Administrative data

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2002
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
comparable to guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2002
Report Date:
2002

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
EPA OPPTS 870.3100 (90-Day Oral Toxicity in Rodents)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent

Test animals

Species:
rat
Strain:
other: Crl:CD(SD)IGS BR
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, NC
- Age at study initiation: approximately 7 weeks
- Weight at study initiation: Males 225 - 309g Females 160 - 214g
- Fasting period before study: No
- Housing: individually in wire mesh cages
- Diet: Certified Rodent Lab Diet 5002, PMI Nutrition Inc., ad libitum except overnight prior to blood collection
- Water: Reverse osmosis treated on-site drinking water ad libitum
- Acclimation period: 15 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22.1 - 22.7
- Humidity (%): 33.3 - 68.1
- Air changes (per hr): 10 - 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: To: 24 October 2000 to 25 January 2001

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: sesame oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: prepared weekly by weighing appropriate amount of test article and adding sufficient vehicle

VEHICLE
- Concentration in vehicle: 0, 0.4, 2, 10, 25/17.5 mg/mL
- Amount of vehicle: 2mL/kg
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Homogeneity and stability established before start of study.
Weekly for first 4 weeks then monthly analysis of formulations for test article concentration during the study
Duration of treatment / exposure:
13 weeks
Frequency of treatment:
5 days/week
Doses / concentrationsopen allclose all
Dose / conc.:
0.8 mg/kg bw/day (actual dose received)
Dose / conc.:
4 mg/kg bw/day (actual dose received)
Dose / conc.:
20 mg/kg bw/day (actual dose received)
Dose / conc.:
50 mg/kg bw/day (actual dose received)
Dose / conc.:
35 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
20 per sex in Control and high dose (50/35 mg/kg/day) groups
10 per sex in intermediate dose groups
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: not stated
Initial high dose of 50 mg/kg/day not tolerated and reduced to 35 mg/kg/day from Day 5.
- Post-exposure recovery period in satellite groups: 28-day recovery period initially included but due to excessive mortality at high dose, no recovery period conducted.

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: at least twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION: Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/animal/day: Yes

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: pre-dose and Week 12
- Dose groups that were examined: All

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Week 4 and 13
- Anaesthetic used for blood collection: No
- Animals fasted: Yes
- How many animals: all
- Parameters checked in table No. 1 were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Week 4 and 13
- Animals fasted: Yes / No / No data
- How many animals: all
- Parameters checked in table No. 2 were examined.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: Week 12
- Dose groups that were examined: all
- Battery of functions tested: sensory activity / grip strength / motor activity ( see table No. 3)

LOCOMOTOR ACTIVITY: Yes
- Time schedule for examinations: Week 12
- Dose groups that were examined: all
- Parameters: ambulatory and total motor activity

OTHER:
BODY TEMPERATURES: recorded from 5 animals/sex/group approx 1 hour after dosing during first 2 weeks of treatment.
Sacrifice and pathology:
GROSS PATHOLOGY: Yes (see table 3)
ORGAN WEIGHT : Yes (see table 3)
HISTOPATHOLOGY: Yes (see table 3)
Statistics:
All analyses were two-tailed for significance levels of 5% and 1%. Body weights, body weight changes, food consumption, clinical pathology values and organ weights were subjected to a one-way analysis of variance (ANOVA) followed by Dunnett's Test if the ANOVA revealed statistical significance (p<0.05). Continuous Functional Observational Battery (FOB) and Locomotor Activity data were analysed using one-way ANOVA. If significant effects were observed at a given timepoint, Dunnett's test was conducted to determine significant treatment differences from the control group. Discontinuous (ordinal or descriptive) FOB data were analysed using Fisher's Exact test to determine significant treatment differences from the control group.

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Twenty-two animals were found dead or killed due to poor clinical condition during the study. One male and 3 females were found dead or killed following administration of 50 mg/kg/day. After reduction in the dose to 35 mg/kg/day, 7 males and 7 females were killed or found dead. At 20 mg/kg/day 1 female was found dead and 1 killed. The majority of animals died during the first 5 weeks of dosing and the deaths were considered test-article related. Two control males were also found dead in weeks 4 and 5 respectively.

Test-article related clinical signs were noted at 50/35 and 20 mg/kg/day and primarily included ataxia, impaired use of hindlimbs, hindlimb tremors, walking on metatarsus, increased salivation and red or yellow material on body surfaces. Other clinical signs noted during the study included prostration, head bobbing, body cool to touch, shallow respiration, lethargy and piloerection.

During detailed clinical observations slight to total impaired mobility was observed at 50/35 mg/kg/day generally throughout the study. Abnormal gait was also noted and included ataxia, dragging hindlimbs, walking on metatarsus and hunched gait.
Mean body temperature was lower for 50/35 mg/kg/day females on Day 2.
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Mean body weight and body weight gain were lower than control for 50/35 mg/kg/day animals and 20 mg/kg/day males generally throughout the study. Overall weight gain was up to 16% lower than control during the dosing period.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Mean food consumption was lower than control for the 50/35 mg/kg/day group generally throughout the study.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
no effects observed
Description (incidence and severity):
No effect of treatment.
Haematological findings:
no effects observed
Description (incidence and severity):
No effect of treatment.
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
Test-article related increases in urea nitrogen were noted at 50/35 and 20 mg/kg/day at weeks 4 (males only) and 13 (males and females). Mean phosphorus levels were increased for 20 mg/kg/day males and 50/35 mg/kg/day animals at both evaluations during the study. Mean potassium levels were increased for 20 mg/kg/day and 50/35 mg/kg/day males during the study and for 50/35 mg/kg/day females in Week 13. Mean cholesterol was lower in 50/35 mg/kg/day males at Week 13.
Urinalysis findings:
not examined
Behaviour (functional findings):
effects observed, treatment-related
Description (incidence and severity):
When evaluating results it should be noted that many of the high dose animals with the most severe hindlimb effects did not survive until the evaluation in week 12.
There were no test-article effects on home cage observations.
An increased number of animals at 50/35 mg/kg/day were noted to have hard and tense muscle tone during the handling observations.
Slightly impaired mobility and abnormal gait was noted for 50/35 mg/kg/day females during the open field observations.
There were no test-article effects on sensory observations.
One female at 50/35 mg/kg/day was noted with exceptionally low hindlimb grip strength during the neuromuscular observations.
There were no remarkable differences apparent between control and treated groups for physiological parameters.
When evaluating results, it should be noted that many of the high dose animals with the most severe hindlimb effects did not survive until the evaluation in week 12.
Overall no remarkable differences between control and treated animals, one female at 50/35 mg/kg/day was noted with the lowest combined motor activity counts.
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Increased mean absolute and relative (to final body weight and brain weight) liver weights were noted at 20 and 50/35 mg/kg/day. Lower seminal vesicle (absolute and relative) weights were also noted for males in these groups and lower prostate (absolute and relative) weights were noted in 50/35 mg/kg/day males.
Gross pathological findings:
no effects observed
Description (incidence and severity):
No findings related to treatment.
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Test-article related microscopic changes were noted in the liver, skeletal muscle and heart and were generally comparable between decedent animals and those surviving to scheduled necropsy.

In the liver, periportal vacuolar change (hepatocytes enlarged by a variably distinct micro- to macro vacuolated cytoplasm) was noted with dose-related incidence in all treated groups. In the absence of correlating clinical chemistry findings, these hepatic changes was considered not to be adverse.

In the skeletal muscle (rectus femoris), minimal to moderate degeneration (hyaline and basophilic cytoplasm, shape and size variations of myocytes, loss of cross striations and increased cellularity) was noted in the 20 and 50/35 mg/kg/day groups.

In the heart, treatment-related or treatment-exacerbated subacute (accumulations of mononuclear cells, primarily macrophages within and surrounding degenerative myocytes) or chronic (mononuclear inflammatory cells and/or fibroplasia and scattered degeneration and necrosis of myocytes) myopathy was noted in 4, 20 and 50/35 mg/kg/day groups. This finding was morphologically similar to spontaneously developing cardiomyopathy typically seen in older rats, however, it was considered to be test-article related in this case.
Histopathological findings: neoplastic:
no effects observed
Details on results:
CLINICAL SIGNS AND MORTALITY:
Twenty-two animals were found dead or killed due to poor clinical condition during the study. One male and 3 females were found dead or killed following administration of 50 mg/kg/day. After reduction in the dose to 35 mg/kg/day, 7 males and 7 females were killed or found dead. At 20 mg/kg/day 1 female was found dead and 1 killed. The majority of animals died during the first 5 weeks of dosing and the deaths were considered test-article related. Two control males were also found dead in weeks 4 and 5 respectively.

Test-article related clinical signs were noted at 50/35 and 20 mg/kg/day and primarily included ataxia, impaired use of hindlimbs, hindlimb tremors, walking on metatarsus, increased salivation and red or yellow material on body surfaces. Other clinical signs noted during the study included prostration, head bobbing, body cool to touch, shallow respiration, lethargy and piloerection.

During detailed clinical observations slight to total impaired mobility was observed at 50/35 mg/kg/day generally throughout the study. Abnormal gait was also noted and included ataxia, dragging hindlimbs, walking on metatarsus and hunched gait.

BODY WEIGHT AND WEIGHT GAIN:
Mean body weight and body weight gain were lower than control for 50/35 mg/kg/day animals and 20 mg/kg/day males generally throughout the study. Overall weight gain was up to 16% lower than control during the dosing period.

FOOD CONSUMPTION:
Mean food consumption was lower than control for the 50/35 mg/kg/day group generally throughout the study.

OPHTHALMOSCOPIC EXAMINATION:
No effect of treatment.

HAEMATOLOGY:
No effect of treatment.

CLINICAL CHEMISTRY
Test-article related increases in urea nitrogen were noted at 50/35 and 20 mg/kg/day at weeks 4 (males only) and 13 (males and females). Mean phosphorus levels were increased for 20 mg/kg/day males and 50/35 mg/kg/day animals at both evaluations during the study. Mean potassium levels were increased for 20 mg/kg/day and 50/35 mg/kg/day males during the study and for 50/35 mg/kg/day females in Week 13. Mean cholesterol was lower in 50/35 mg/kg/day males at Week 13.

NEUROBEHAVIOUR:
When evaluating results it should be noted that many of the high dose animals with the most severe hindlimb effects did not survive until the evaluation in week 12.
There were no test-article effects on home cage observations.
An increased number of animals at 50/35 mg/kg/day were noted to have hard and tense muscle tone during the handling observations.
Slightly impaired mobility and abnormal gait was noted for 50/35 mg/kg/day females during the open field observations.
There were no test-article effects on sensory observations.
One female at 50/35 mg/kg/day was noted with exceptionally low hindlimb grip strength during the neuromuscular observations.
There were no remarkable differences apparent between control and treated groups for physiological parameters.

LOCOMOTOR ACTIVITY:
When evaluating results, it should be noted that many of the high dose animals with the most severe hindlimb effects did not survive until the evaluation in week 12.
Overall no remarkable differences between control and treated animals, one female at 50/35 mg/kg/day was noted with the lowest combined motor activity counts.

ORGAN WEIGHTS:
Increased mean absolute and relative (to final body weight and brain weight) liver weights were noted at 20 and 50/35 mg/kg/day. Lower seminal vesicle (absolute and relative) weights were also noted for males in these groups and lower prostate (absolute and relative) weights were noted in 50/35 mg/kg/day males.

GROSS PATHOLOGY:
No findings related to treatment.

HISTOPATHOLOGY:
Test-article related microscopic changes were noted in the liver, skeletal muscle and heart and were generally comparable between decedent animals and those surviving to scheduled necropsy.

In the liver, periportal vacuolar change (hepatocytes enlarged by a variably distinct micro- to macro vacuolated cytoplasm) was noted with dose-related incidence in all treated groups. In the absence of correlating clinical chemistry findings, these hepatic changes was considered not to be adverse.

In the skeletal muscle (rectus femoris), minimal to moderate degeneration (hyaline and basophilic cytoplasm, shape and size variations of myocytes, loss of cross striations and increased cellularity) was noted in the 20 and 50/35 mg/kg/day groups.

In the heart, treatment-related or treatment-exacerbated subacute (accumulations of mononuclear cells, primarily macrophages within and surrounding degenerative myocytes) or chronic (mononuclear inflammatory cells and/or fibroplasia and scattered degeneration and necrosis of myocytes) myopathy was noted in 4, 20 and 50/35 mg/kg/day groups. This finding was morphologically similar to spontaneously developing cardiomyopathy typically seen in older rats, however, it was considered to be test-article related in this case.

BODY TEMPERATURE:
Mean body temperature was lower for 50/35 mg/kg/day females on Day 2.

Effect levels

Key result
Dose descriptor:
NOAEL
Effect level:
0.8 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: see 'Remark'

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

Table 1 - survival

Dose
(mg/kg/day)

Control

0.8

4

20

50/35

Males

Number in group

20

10

10

10

20

Found dead

2

0

0

0

4

Killed

0

0

0

0

4

Survived until scheduled termination

18

10

10

10

12

Females

Number in group

20

10

10

10

20

Found dead

0

0

0

1

4

Killed

0

0

0

1

6

Survived until scheduled termination

20

10

10

8

10

Applicant's summary and conclusion

Conclusions:
Oral administration of trifluoropropylmethylcyclotrisiloxane (TFPMCT) to rats at 0.8, 4, 20 or 50/35 mg/kg/day for 13 weeks was not tolerated at the highest dose and resulted in a number of mortalities, 2 animals also died at 20 mg/kg/day. Pathological changes were noted in the liver, heart and skeletal muscle and several clinical findings were noted consistent with the skeletal muscle toxicity. Generally, effects were primarily noted at 20 or 50 mg/kg/day and also included body weight, food consumption, clinical chemistry and organ weight effects.

Based on mortality at 20 or 50/35 mg/kg/day, clinical signs, body weight, food consumption, clinical chemistry, FOB and organ weight effects at 20 and/or 50/35 mg/kg/day and pathology findings in the skeletal muscle and heart at 4, 20 or 50/35 mg/kg/day, the NOAEL was considered to be 0.8 mg/kg/day.