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Repeated dose toxicity: oral

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Administrative data

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

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2003
Report date:
2003

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
Remarks:
The study was conducted according to the test guidelines in effect at the time of study conduct.
Qualifier:
according to guideline
Guideline:
other: 28-Day repeated administration toxicity test to use mammals (Partial ammendment-Novel chemical substance testing method [Environ. Protect. Notification 700, Notification 1039 of Pharmaceutical Affairs Bureau & MITI Basic Industry Bureau Notification 1014)
Deviations:
no
Remarks:
The study was conducted according to the test guidelines in effect at the time of study conduct.
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Carboxymethyldimethyl-3-[[(3,3,4,4,5,5,6,6,7,7,8,8,8-tridecafluorooctyl)sulphonyl]amino]propylammonium hydroxide
EC Number:
252-046-8
EC Name:
Carboxymethyldimethyl-3-[[(3,3,4,4,5,5,6,6,7,7,8,8,8-tridecafluorooctyl)sulphonyl]amino]propylammonium hydroxide
Cas Number:
34455-29-3
Molecular formula:
C15H19F13N2O4S
IUPAC Name:
carboxymethyldimethyl-3-[[(3,3,4,4,5,5,6,6,7,7,8,8,8-tridecafluorooctyl)sulphonyl]amino]propylammonium hydroxide
Details on test material:
- Purity: 99.6%

Test animals

Species:
rat
Strain:
other: Crj:CD(SD) IGS
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Nippon Charles River K.K.
- Age at study initiation: 5 weeks
- Weight at study initiation: 141.7-159.9 g (males); 115.5-132.9 g (females)
- Fasting period before study:
- Housing: Prior to grouping, the animals were kept 5 head/cage in stainless steel and metal mesh floor cages. After grouping, the animals were individually kept in stainless steel and metal mesh cages.
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: Not reported

ENVIRONMENTAL CONDITIONS
- Temperature (°C): targeted at 21-25°C; actual 21.9-23.9°C
- Humidity (%): targeted at 40-70%; actual 54.6-71.8% (on day 2 of the recovery term, the function of the air conditioner was suspended for 8 minutes due to being hit by lightning causing the humidity to rise [maximum humidity of 71.8%]. No abnormality was observed in the animals and the incident was considered not to affect the result of the study.)
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12-hr interval

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
The test substance was accurately weighed and suspended in pure water by adding a small amount of pure water at a time into an agate mortar to prepare 0.1, 0.4, 2.0 and 10.0 w/v% solutions. The test substance preparation was carried out once a week.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The stability of the test substance was confirmed by measuring its infrared absorption spectrum before the start and after the end of the administration term. The infrared absorption spectrum measurements were carried out by using a Fourier transform infrared spectrophotometer in the range of 4000 cm-1 to 400 cm-1. Before the start of the administration, the identification was carried out by comparison with a spectrum provided by the test requester, and in the measurement carried out after completing the administration term, the spectrum before the administration was used as a comparison.

Stability and uniformity of the test substance formulations were carried out. As a result, the 10.0 and 0.05 w/v% solutions prepared were confirmed to be stable in a cool and dark place for 7 days. Furthermore, the samples were found to be homogeneous. For the homogeneity of the test substance solutions prepared, immediately after preparation, the 10.0 w/v% and 0.05 w/v% solutions were sampled from top, middle and bottom layers with n = 3 for each layer. The samples were diluted with methanol, the test substance concentration was measured by using high performance liquid chromatography (HPLC), and the homogeneity was confirmed from the relative standard deviation of the test substance concentrations at respective sampling sites. For the stability confirmation, the 10.0 w/v% and 0.05 w/v% solutions were stored in a cool and dark place, and 3 samples from the middle layer were collected after 3 days and 7 days. The samples were diluted with methanol, the test substance concentration was measured by using high performance liquid chromatography (HPLC), and the stability was confirmed by finding any change from the concentrations immediately after preparation (results of homogeneity test).

Analysis conditions
Instrument used (HPLC)
Measurement conditions:
Column: Shodex Asahipak GF-310HQ 7.6 mm I.D. x 300 mm
Column temperature: 40°C
Mobile phase: methanol
Flow rate: 1 mL/min
Amount injected: 50 µL
Duration of treatment / exposure:
28 days
14-day recovery for 0 and 1000 mg/kg
Frequency of treatment:
Daily
Doses / concentrationsopen allclose all
Dose / conc.:
10 mg/kg bw/day (actual dose received)
Dose / conc.:
40 mg/kg bw/day (actual dose received)
Dose / conc.:
200 mg/kg bw/day (actual dose received)
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
6/sex/dose at 10, 40, and 200 mg/kg
12/sex/dose at 0 and 1000 mg/kg
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: A 14-day repeated oral administration toxicity test was carried out at doses of 50, 250, 500 and 1000 mg/kg/day. As a result, there were changes in the general condition observed in the 250 mg/kg group that were suspected to be attributable to the test substance. Therefore, in the present study, the high dose was set at 1000 mg/kg/day followed by 200, 40, and 10 mg/kg/day. Recovery groups were set for 0 (control) and 1000 mg/kg/day.

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: 3 times/day before substance administration (in the morning), during and after substance administration and in the afternoon every day from day 1 to day 28. During recovery, observation was carried out once a day in the morning.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Symptoms were summarized every week.

BODY WEIGHT: Yes
- Time schedule for examinations: Day -2 (at the time of grouping), days 1, 3, 8, 12, 17, 21, 26, and 28 during administration, and day 1, 5, 10, and 14 of recovery.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Time schedule for examinations: Once before substance administration, on days 3, 8, 15, 22, and 28 during administration, and days 4, 8, and 14 of recovery

FOOD EFFICIENCY: No

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

OPHTHALMOSCOPIC EXAMINATION: No data

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Day 28 of administration (excluding recovery animals) and day 14 of recovery
- Anaesthetic used for blood collection: Yes (ether)
- Animals fasted: Yes (16-20 hours)
- How many animals: all
- Parameters checked in table No. 1 were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Day 28 of administration (excluding recovery animals) and day 14 of recovery
- Anaesthetic used for blood collection: Yes (ether)
- Animals fasted: Yes (16-20 hours)
- How many animals: all
- Parameters checked in table No. 2 were examined.

URINALYSIS: Yes
- Time schedule for collection of urine: Day 28 of administration (excluding recovery animals) and day 14 of recovery
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes
- Parameters examined: Amount and colouration, pH, protein, ketone body, bilirubin, occult blood, glucose, urobilinogen using test paper (N-Multistix®)

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes (see table 3)
HISTOPATHOLOGY: Yes (see table 4)

Statistics:
The results on bodyweight (except at the time of body removal), feed intake. Haematological tests, clinical chemistry tests, amount of urine and organ weights were tested for equality of variance by the Bartlett method, and if there was equal variance observed at a 5% significance level, a one-way analysis of variance was carried out. If the results of analysis of variance showed a significant difference, a test by the Dunnett method was carried out between the control group and respective administration groups. If no equal variance was observed, a Kruskal-Wallis test was carried out, and if there was a significant difference observed, a test was carried out between the control group and respective administration groups by using the non-parametric Dunnett method.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Description (incidence and severity):
Both males and females at 200 mg/kg or higher showed soft stool considered to be caused by the test substance administration. However, no effects were observed on the results of pathological tests for changes in the digestive tract, etc. During the recovery period, soft stool was observed in both males and females at 1000 mg/kg, but both males and females showed no soft stool on day 4 indicating that the effect was reversible.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Male body weights were reduced at 1000 mg/kg from day 12 to 28 of the administration period. Male body weight was reduced at 1000 mg/kg from day 1 to 14 of the recovery period, but gains were similar to those observed in the controls. Therefore, the reductions in body weight were considered reversible.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
An increase in feed consumption was observed in females at 1000 mg/kg on day 8 of recovery. No other abnormalities were observed.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Description (incidence and severity):
Males at 200 mg/kg had shortened activated partial thromboplastin time at the end of administration. No other abnormalities were observed.
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
Reduced albumin was observed in males at 200 mg/kg or higher, but the changes were all fluctuations within the background value range of the Laboratory and no dose-dependency of the results was found. Therefore, the changes were not considered related to administration of the test substance.
Urinalysis findings:
no effects observed
Description (incidence and severity):
Increased amount of urine was observed in 1000 mg/kg males at the end of the recovery period. No other abnormalities were observed.
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Increased relative liver weight was observed in 1000 mg/kg females at the end of the administration period. Increased relative weights of testis and brain were observed in 1000 mg/kg males at the end of the recovery period. No other abnormalities were observed.
Gross pathological findings:
no effects observed
Description (incidence and severity):
At the end of the administration period, pyelectasia of the kidney was observed in 1/6 males in the control and the 40 mg/kg groups; and whitened section of the spleen was observed in 1/6 males at 200 mg/kg. At the end of the recovery period, a node in the spleen was observed in 1/6 males at 1000 mg/kg. No abnormalities were observed in females.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Centriloblular hepatic cell hypertrophy was observed in 3/6 males and a single cell necrosis in 4/6 males at 1000 mg/kg at the end of administration. Pyelectasia of the kidney was observed in males at 0 (1/6) and 40 mg/kg (1/6). A small granuloma in the liver (1/6) and local necrosis in the spleen (1/1) was observed in males at 200 mg/kg. At the end of administration, only a single occurrence cyst in the medulla of the kidney was observed in one female at 0 mg/kg. At the end of the recovery period in males, a small granuloma of the liver (1/6) at 0 mg/kg and follicular hypertrophy of the spleen at 1000 mg/kg (1/1) was observed.
Histopathological findings: neoplastic:
no effects observed

Effect levels

open allclose all
Dose descriptor:
NOEL
Effect level:
40 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: soft stool in males and females at ≥200 mg/kg
Key result
Dose descriptor:
NOAEL
Effect level:
200 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
other: hepatic single cell necrosis and hypertrophy, which showed recovery, in males at 1000 mg/kg

Target system / organ toxicity

Key result
Critical effects observed:
no
Lowest effective dose / conc.:
1 000
System:
hepatobiliary
Organ:
liver
Treatment related:
yes
Dose response relationship:
no
Relevant for humans:
not specified

Applicant's summary and conclusion

Conclusions:
NOEL: 40 mg/kg (based on clinical signs of soft stool at ≥200 mg/kg)
NOAEL: 200 mg/kg (based on hepatic single cell necrosis and hypertrophy, which showed recovery, in males at 1000 mg/kg)

This study and the conclusions which are drawn from it fulfil the quality criteria (validity, reliability, repeatability).
Executive summary:

A 28-day repeated oral administration toxicity test and was carried in male and female rats at doses of 0, 10, 40, 200, or 1000 mg/kg/day. A 14-day recovery period was included at 0 and 1000 mg/kg/day. 

No mortality attributable to the test substance was observed. There were no toxicological effects of test substance administration with respect to haematology, clinical chemistry, urinalysis, or gross pathology. Males and females at ≥200 mg/kg had soft stool, which was considered to be test substance related. In addition, males at 1000 mg/kg had reduced body weights from day 12 to day 28. Increased relative liver weight was observed in females at 1000 mg/kg. Centrilobular hepatic cell hypertrophy and single cell necrosis in the liver were observed in males at 1000 mg/kg. In the recovery test, all changes observed during the administration term and at the end of the administration terms showed recovery. The results as described above suggested test substance-related effects of soft stool, decreases in bodyweight, and liver changes, which were all considered reversible. The NOEL under the conditions of this study was 40 mg/kg/day based on the observation of soft stool in males and females at doses of ≥200 mg/kg/day.