Registration Dossier

Administrative data

Endpoint:
short-term repeated dose toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
08 Feb - 12 Jul 2005
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

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 410 (Repeated Dose Dermal Toxicity: 21/28-Day Study)
Version / remarks:
adopted in 1981
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.9 (Repeated Dose (28 Days) Toxicity (Dermal))
Version / remarks:
adopted in 1992
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.3200 (Repeated Dose Dermal Toxicity -21/28 Days)
Version / remarks:
adopted in 1998
Qualifier:
according to guideline
Guideline:
other: M.A.F.F. in Japan 12 NohSan N° 8147
Version / remarks:
adopted in 2001
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
(5-hydroxy-1,3-dimethyl-1H-pyrazol-4-yl)[2-(methylsulfonyl)-4-(trifluoromethyl)phenyl]methanone
EC Number:
609-256-3
Cas Number:
365400-11-9
Molecular formula:
C14H13F3N2O4S
IUPAC Name:
(5-hydroxy-1,3-dimethyl-1H-pyrazol-4-yl)[2-(methylsulfonyl)-4-(trifluoromethyl)phenyl]methanone
Test material form:
solid: crystalline

Test animals

Species:
rat
Strain:
Wistar
Remarks:
HsdCpb:WU
Details on species / strain selection:
The rat is the recommended species for repeat-dose dermal toxicity studies. Animals of this strain have been used at the Bayer HealthCare AG for toxicological studies for many years. Historical data on their physiology and spontaneous alterations is available.
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Harlan Winkelmann GmbH Experimental Animal Breeders in Borchen, Germany
- Females nulliparous and non-pregnant: yes
- Age at study initiation: males: 9 - 10 weeks, females: 13 - 14 weeks
- Weight at study initiation: males: 222 - 262 g (mean 244 g), females: 202 - 230 g (mean 213 g)
- Fasting period before study: no
- Housing: individually in Type IIIh cages on wood shavings
- Diet: PROVIMI KLIBA® 3883.0.15 (Provimi Kliba SA, Kaiseraugst, Switzerland), ad libitum
- Water: tap water, ad libitum
- Acclimation period: at least seven days

DETAILS OF FOOD AND WATER QUALITY: analysis was performed

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 24
- Humidity (%): 50 - 60
- Air changes (per hr): more than 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 08 Feb 2005 To: 09 Mar 2005

Administration / exposure

Type of coverage:
semiocclusive
Vehicle:
unchanged (no vehicle)
Remarks:
The test item was applied as undiluted solid onto a wet gauze pad.
Details on exposure:
TEST SITE
- Area of exposure: back and flank region, 6.0 x 5.0 cm = 30.0 cm²
- % coverage: greater than 10% of body surface area
- Type of wrap if used: the test item was applied to the moist gauze-layer of a "Cutiplast steril®" patch and secured in place with using Peha-Haft® placed on the rat's back; the gauze-layer was secured in place using "Peha-Haft®" cohesive stretch tape (approx. 8 x 23 cm).
- Time intervals for shavings: one day before the start of treatment and twice weekly afterwards

REMOVAL OF TEST SUBSTANCE
- Washing (if done): test item was removed with soap and water
- Time after start of exposure: 6 h

TEST MATERIAL
- Amount(s) applied: 10, 100 and 1000 mg/kg bw/day;
males reveived 9.6 - 12.8, 7.3 - 10.8 and 15.5 - 19.8 mg/cm²/day; females received 8.0 - 8.4, 5.7 - 9.5 and 15.0 - 16.1 mg/cm²/day
- For solids, paste formed: no

USE OF RESTRAINERS FOR PREVENTING INGESTION: yes (,,Lomir Biomedical Inc." rat jacket)
Analytical verification of doses or concentrations:
no
Remarks:
(No dose formulation preparation or analysis was performed during the study, as the test item powder was applied to the rat skin without using any vehicle (only moistened with water)).
Details on analytical verification of doses or concentrations:
not applicable
Duration of treatment / exposure:
study period 28 days (males received 22 applications, females 23)
Frequency of treatment:
first three weeks: 5 days/week; thereafter: 7 days/week
Doses / concentrationsopen allclose all
Dose / conc.:
10 mg/kg bw/day
Dose / conc.:
100 mg/kg bw/day
Dose / conc.:
1 000 mg/kg bw/day
No. of animals per sex per dose:
10
Control animals:
yes, sham-exposed
Details on study design:
- Dose selection rationale: Dose levels were selected and determined by the sponsor according to results obtained in previous studies.

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Inspections on mortality and morbidity of the animals were done twice daily and on weekends / on public holidays once a day.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: A careful clinical examination including observation outside the home cage in a standard arena (OFO) was made once prior to treatment and daily (careful clinical examination) or weekly (OFO) thereafter up to necropsies in all animals.

DERMAL IRRITATION (if dermal study): Yes
- Time schedule for examinations: The shaved skin areas were examined before starting the study and each day of treatment during the study.

BODY WEIGHT: Yes
- Time schedule for examinations: The animals were weighed before the study was started and at the beginning of each study week.

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

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: No

WATER CONSUMPTION: Yes
- Time schedule for examinations: Individual water intake was determined once weekly from the start of the study.

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: Ophthalmological examinations were performed before the start of the study and at the end of the treatment period.
- Dose groups that were examined: all (pre-treatment), groups 1 (control) and 4 (high dose) at the end of the treatment period

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Blood sampling was performed before the start of the study and towards the end of the treatment period (males day 25, females day 28).
- Anaesthetic used for blood collection: Yes (CO2/O2)
- Animals fasted: No
- How many animals: all
- Parameters examined: leucocyte count, differential blood count, erythrocytes, hemoglobin, hematocrit, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, mean corpuscular volume, thrombocytes, reticulocytes, hepatoquick

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Blood sampling was performed before the start of the study and towards the end of the treatment period (males day 25, females day 28).
- Animals fasted: No
- How many animals: all
- Parameters examined: alanine aminotransferse activity, albumin concentration, alkaline phosphatase activity, aspartate aminotransferase activity, total bilirubin concentration, cholesterol concentration, creatinine concentration, calcium concentration, chloride concentration, glucose concentration, potassium concentration, sodium concentration, inorganic phosphate concentration, total protein concentration, triglycerides, urea concentration

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes, all animals alive at scheduled necropsy as well as those killed in moribund state

HISTOPATHOLOGY: Yes
- adrenal glands, aorta, brain (cerebrum, cerebellum, brain stem), epididymides, esophagus, eyes, eyelids, exorbital lacrimal glands, femur, harderian glands, head: nasal cavity, heart, intestine (incl. Peyer's patches): duodenum, jejunum, iileum, caecum, colon, rectum, remaining intestine, kidneys, larynx, liver, lungs, lymph nodes: mandibular and mesenteric, optic nerves, ovaries, oviducts, physical identifier, pancreas, pharynx, pituitary gland, prostate, salivary glands (parotid, submandibular, sublingual), sciatic nerve, seminal vesicles (incl. coagulation glands), skeletal muscle (tigh), skin (mammary region), spinal cord (cervical, thoracic, lumbar), spleen, sternum, stomach (forestomach and glandular stomach), sestes, thymus, thyroid glands (with parathyroids), tongue, trachea, ureters, urethra, urinary bladder, uterus (with cervix), vagina, Zymbal's glands, organs and tissues with macroscopic findings, skin - treated, skin - untreated

ORGAN WEIGHTS: Yes
- adrenal glands, kidneys, liver, brain, testes, epididymides, heart, ovaries, spleen, thyroid gland (in fixed state), thymus, uterus
Statistics:
As basis for statistical evaluation of numerical data SAS 6.12 was used. The results for the groups that received the test compound were compared with those for the control group. The statistical evaluation of data related to laboratory investigations, body and organ weights as well as food and water intake is performed using SAS® routines.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Dermal irritation:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
no effects observed
Haematological findings:
effects observed, non-treatment-related
Description (incidence and severity):
In mid-dose males a statistically significantly higher MCHC value was noted (320 g/L Ery vs. 314 in the control). In mid-dose females a statistically significantly higher MCH value was noted (17.4 pg vs. 16.9 in the control). Both differences were only mimimal in degree and showed no dependency. Therefore these changes were considered to be not treatment-related.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
The activities of aspartate aminotransferase were decreased at 10 mg/kg bw/day and above and of alkaline phosphatase at 100 mg/kg bw/day and above in both sexes.
The concentrations of cholesterol, triglycerides, total protein and albumin were higher, and the concentrations of creatinine lower in males at 1000 mg/kg bw/day. In females, the concentration of bilirubin was higher at 100 and 1000 mg/kg bw/day.
The concentration of sodium was decreased at 100 mg/kg bw/day and above in both sexes.
Calcium was increased at 1000 mg/kg in females, and inorganic phosphate decreased in males at 10 mg/kg bw/day and above.
All these partly statistically significant differences were either of a minimal degree, and/or not dose-related, and/or were observed only in one sex, and/or were all in the range of historical values. Therefore, they were considered as not treatment-related.
Urinalysis findings:
not examined
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):
Absolute and relative liver weights were statistically significantly increased at 1000 mg/kg bw/day in both sexes.

Absolute mean liver weight was statistically significantly increased in females at 10 mg/kg bw/day. However, relative mean liver weight of these animals was similar to the control group. In the absence of liver weight effects at 100 mg/kg bw/day this finding was considered incidental in nature.
Gross pathological findings:
no effects observed
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Locally, no evidence of any findings was detected at the application site of the skin.
In the pancreas, focal degeneration was detected at 100 mg/kg bw/day and above in both sexes.
In the thyroid gland, follicular cell hypertrophy was detected in both sexes in all dose groups, including controls. However, an increased incidence occurred in males at 1000 mg/kg bw/day (5-5-4-9 affected animals with increasing doses). Furthermore, colloid alteration of the thyroid was detected with an increased incidence in males (1-4-6-10 affected animals with increasing doses). The incidence occurred together with an increasing grading in the upper dose groups.
Slight hepatocellular hypertrophy was observed at 1000 mg/kg bw/day, in males.
Hypertrophy of the pars distalis of pituitary was observed at 1000 mg/kg bw/day in males.
Histopathological findings: neoplastic:
no effects observed
Other effects:
not examined
Details on results:
Thyroid findings (increased weight, histopathological changes comprising changes in colloid, follicular cell hypertrophy and pigment deposition in the follicular epithelium) are considered a non-adverse and rat specific phenomenon. No changes of the thyroid were noted in either mice or dogs, the other two species in which repeated-dose studies with histopathological examination of the thyroid were conducted with the test substance. As the test substance through inhibition of the HPPDase enzyme increases plasma tyrosine concentration in the rat, it is quite possible that some of this increased tyrosine is taken up by the thyroid and stored in the colloid, either as free tyrosine or through either increasing the synthesis of thyroglobulin or altering its composition in terms of number of tyrosine residues per thyroglobulin molecule.
In the absence of signs of altered thyroid functions (such as effects on body weights, fertility and gestation indices or effects on offspring performance in the developmental neurotoxicity study), the observed morphological changes are considered to be non-adverse.

Effect levels

open allclose all
Key result
Dose descriptor:
NOAEL
Remarks:
local
Effect level:
1 000 mg/kg bw/day
Based on:
test mat.
Remarks:
equivalent to 15.0 mg/m2
Sex:
male/female
Basis for effect level:
other: No dermal irritation observed up to the highest dose tested.
Key result
Dose descriptor:
NOAEL
Effect level:
10 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
histopathology: non-neoplastic
organ weights and organ / body weight ratios

Target system / organ toxicity

open allclose all
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
100 mg/kg bw/day
System:
gastrointestinal tract
Organ:
pancreas
Treatment related:
yes
Dose response relationship:
yes
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
1 000 mg/kg bw/day
System:
endocrine system
Organ:
pituitary gland
Treatment related:
yes
Dose response relationship:
yes
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
1 000 mg/kg bw/day
System:
hepatobiliary
Organ:
liver
Treatment related:
yes
Dose response relationship:
yes
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
1 000 mg/kg bw/day
System:
endocrine system
Organ:
thyroid gland
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
no

Any other information on results incl. tables

Table 2: Mean liver weights on Day 28

 

Dose groups (mg kg bw/day)

 

control

10

100

1000

Males

Absolute (g)

14.42

13.90

13.71

16.43*

Relative to Body Weight (%)

4.23

4.48

4.33

4.98**

Females

Absolute (g)

8.49

9.20*

8.50

9.45**

Relative to Body Weight (%)

3.75

3.94

3.75

4.19**

* = p<0.05, ** = p< 0.01 (Dunnett test)

Applicant's summary and conclusion

Conclusions:
Based on the effects observed at 100 mg/kg bw/day, the substance is classified "STOT RE 2, H373, dermal, gastro-intestinal".