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Toxicological information

Repeated dose toxicity: oral

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Administrative data

Endpoint:
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:
1999
Report date:
1999

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 452 (Chronic Toxicity Studies)
Qualifier:
according to guideline
Guideline:
EU Method B.30 (Chronic Toxicity Studies)
Qualifier:
according to guideline
Guideline:
EPA OPP 83-1 (Chronic Toxicity)
Qualifier:
according to guideline
Guideline:
other: Japanese Agricultural Chemical Laws and Regulations Japan (ii) (1985) Society of Agricultural Chemical Industry
GLP compliance:
yes

Test material

Constituent 1
Chemical structure
Reference substance name:
methyl (2E)-3-methoxy-2-[2-({[6-(trifluoromethyl)pyridin-2-yl]oxy}methyl)phenyl]acrylate
EC Number:
601-478-9
Cas Number:
117428-22-5
Molecular formula:
C18H16F3NO4
IUPAC Name:
methyl (2E)-3-methoxy-2-[2-({[6-(trifluoromethyl)pyridin-2-yl]oxy}methyl)phenyl]acrylate
Test material form:
solid
Details on test material:
Purity: 93.3-99.8 (see individual test record for specific details)
Specific details on test material used for the study:
Purity: 94.4% (w/w)

Test animals

Species:
dog
Strain:
Beagle
Details on species / strain selection:
The dog is the preferred non-rodent species for regulatory studies and the Alderley Park beagle was used because of the substantial background data available for this strain, in the Laboratory, relating to studies of this type.
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Convential Animal Breeding Unit, Zeneca Pharmaceuticals, Alderley Park, Macclesfield, Cheshire UK
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 16-24 weeks old on delivery
- Weight at study initiation: not provided
- Fasting period before study: no
- Housing: On arrival, dogs were housed by treatment group (sexes separately) in indoor pens. The pens have a sleeping platform with heated floor underneath and interlinking gates which enable the dogs to be separated for feeding. Dogs received exercise (while on a lead) for all routine procedures (weighing) and were allowed weekly access to a free exercise area, on a group and sex basis.
- Diet: 350 g for males and 300 g for females of the appropriate experimental diet/day
- Water: ad libitum
- Acclimation period: 4-5 weeks

ENVIRONMENTAL CONDITIONS
- Temperature: 19±4°C
- Humidity: 40-70%
- Air changes (per hr): Approximately 15 changes/hour
- Photoperiod (hrs dark / hrs light): Artificial giving 12 hours light, 12 hours dark

Administration / exposure

Route of administration:
oral: feed
Details on route of administration:
The dietary route was chosen for administration of the test substance as this represents a possible route of exposure in humans and other mammalian species.
Vehicle:
unchanged (no vehicle)
Details on oral exposure:
DIET PREPARATION
- Rate of preparation of diet (frequency): The experimental diets were prepared in 60 kg batches
- Mixing appropriate amounts: A 1 kg premix was prepared by addition of the test substance to a small amount of LABORATORY DIET A, whilst mixing in a automatic pestle and mortar, and then adding the remaining diet to make up the 1 kg premix. This was then mixed with 59 kg of diet in a blender. Water was added to the batch of diet at a rate of approximately 100 mL/kg diet and the diets mixed twice for 6 minutes in a Hobart mixer. The diet was then pelleted, using a Laboratory Pellet Mill Feeder.
- Storage temperature of food: Room temperature
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples were extracted with acetonitrile
Method: HPLC
HPLC Conditions
- Pump: 600 Series (Waters)
- Mobile phase: Acetonitrile (65% v/v); Water (35% v/v); Phosphoric acid (0.1% v/v)
- Flow rate: 1.5 mL/min
- Detector wavelength: 245 nm
- Column: 25cm x 4.6mm ID Hypersil BDS (Shandon) or 15cm x 3.9mm ID Symmetry C18 (Waters)
- Column temperature: Ambient
- Sample introduction: 717 plus (Waters)
- Injection volume: 20 µL
- Limit of detection: 0.2 µg/mL (corresponding to 2 ppm)
Mean measured concentrations achieved: 47.0, 140, and 485 ppm for 50, 150 and 500 ppm groups respectively. The overall mean concentrations were within 7% of target.
Homogenity and chemical stability were determined to be satisfactory.
Duration of treatment / exposure:
1 year
Frequency of treatment:
Daily
Doses / concentrationsopen allclose all
Dose / conc.:
50 ppm
Dose / conc.:
150 ppm
Dose / conc.:
500 ppm
No. of animals per sex per dose:
4/gender/dose
Control animals:
yes, plain diet
Details on study design:
- Dose selection rationale: The dose levels selected for this study were based on the results of a previous study in the dog carried out in the test facility.

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Weekly

BODY WEIGHT: Yes
- Time schedule for examinations: Weekly

FOOD CONSUMPTION AND COMPOUND INTAKE:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes
- Time schedule for examinations: Food residues were recorded daily, approximately 4 hours after feeding and any residual food was discarded. These measurements were made for at least 2 weeks pre-study and throughout the treatment period. Food consumption was calculated, at weekly intervals, as a mean value for each dog.

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

OPHTHALMOSCOPIC EXAMINATION: Yes

HAEMATOLOGY: Yes
- Time schedule for collection of blood: weeks -1, 13, 26, and prior to termination
- Parameters examined: red blood cell count, haemoglobin, haematocrit, mean cell volume, mean cell haemoglobin, mean cell haemoglobin concentration, total white cell count (WBC), differential WBC count, platelet count

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: weeks -1, 13, 26, and prior to termination
- Parameters examined: alanine aminotransferase activity, albumin, alkaline phosphatase activity, aspartate aminotransferase activity, calcium, chloride, creatinine, creatine kinase activity, gamma-glutamyl transferase activity, glucose, phosphorus (as phosphate), potassium, sodium, total bilirubin, cholesterol, total protein, urea

URINALYSIS: Yes
- Time schedule for collection of urine: weeks -1, 13, 26, and prior to termination
- Parameters examined: volume, appearance, colour (only if abnormal), specific gravity, pH, glucose, ketones, bilirubin, protein, blood. In addition, each urine sample was centrifuged and the sediment stained and examined microscopically to identify components.

NEUROBEHAVIOURAL EXAMINATION: No

IMMUNOLOGY: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes. All animals were subjected to a full examination post mortem, which involved an external observation and a careful examination of all internal organs and structure. The following organs were removed, trimmed free of extraneous tissue and weighed: adrenal glands, brain, kidneys, liver (without gall bladder), ovaries, testes, thyroid glands with parathyroid.

HISTOPATHOLOGY: Yes (see table 1)
Statistics:
All data were evaluated using analysis of variance and/or analysis of covariance for each specified parameter using the GLM procedure in SAS.

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
There was an increased incidence of thin appearance for females receiving 500 ppm. The only other clinical sign that was considered possibly to be related to treatment was an increased incidence of reddened gums for males receiving 500 ppm. There was an increased incidence of reddened ears in males receiving 500 ppm. Reddened ears were observed at all dose levels including controls and are considered to be incidental to administration of the test substance. There were no clinical signs associated with administration of 50 or 150 ppm.

The only gastrointestinal finding that was considered to be related to treatment was a slightly increased incidence of fluid faeces in males receiving 500 ppm.

None of the clinical signs related to treatment were considered to be of toxicoligical significance.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
A treatment-related reduction in body weight was seen for males and females receiving 500 ppm. There were no effects on body weights at 50 or 150 ppm.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Treatment-related effects on food consumption was seen for males and females receiving 500 ppm. There were no treatment-related effects on food consumption at 50 or 150 ppm.
Ophthalmological findings:
no effects observed
Description (incidence and severity):
There were no veterinary or ophthalmoscopy findings that were considered related to treatment.
Haematological findings:
effects observed, non-treatment-related
Description (incidence and severity):
Minor variations in haematology parameters were seen at isolated time points but these were small in magnitude and/or showed no dose response relationship and were considered to be incidental to treatment.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
Minor variations in blood chemistry parameters were seen at isolated time points but these were small in magnitude and/or showed no dose response relationship and were considered to be incidental to treatment.
Urinalysis findings:
effects observed, non-treatment-related
Description (incidence and severity):
Minor variations in urine clinical chemistry parameters were seen at isolated time points but these were small in magnitude and/or showed no dose response relationship and were considered to be incidental to treatment.
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
Group mean thyroid weights, adjusted for terminal body weight, for females receiving 500 ppm were higher than concurrent controls. The control group mean value was low compared to historical control group mean range. Whilst a relationship to treatment cannot be entirely ruled out, in the absence of any associated treatment-related histopathological changes the increase is considered to be of no toxicological significance.

Group mean kidney weights for males receiving 500 ppm were lower than concurrent controls but the reduction was not statistically significant after adjustment for terminal body weight and is considered to be incidental to treatment with the test substance. Group mean brain weights, for females receiving 500 ppm were slightly higher than concurrent controls but the increase did not attain statistical significance and is considered to be incidental to administration of the test substance.
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
Slight dilatation of the lateral ventricles of the brain was observed macroscopically in one animal of each sex receiving 500 ppm and in one male receiving 150 ppm but on histopathological examination this was considered to be within normal limits. The small number of macroscopic changes observed in other tissues were considered to be incidental to administration of the test substance.
Histopathological findings: non-neoplastic:
effects observed, non-treatment-related
Description (incidence and severity):
A number of minor changes were observed but all were considered to be incidental to administration of the test substance.

Effect levels

Key result
Dose descriptor:
NOAEL
Effect level:
150 ppm
Sex:
male/female
Remarks on result:
other: clinical observations, body weight reduction, food consumption effects at 500 ppm

Target system / organ toxicity

Key result
Critical effects observed:
no

Any other information on results incl. tables

Dose rates were calculated in terms of mg/kg/day. Mean values for males were: 1.6, 4.8, and 16.1 mg/kg/day for the 50, 150, and 500 ppm groups respectively. Mean values for females were 1.6, 4.6, and 15.7 mg/kg/day for the 50, 150, and 500 ppm groups respectively

Applicant's summary and conclusion

Conclusions:
Administration of 500 ppm in the diet for 52 weeks resulted in reduced growth and reduced food consumption in males and females. This dose was equivalent to an overall mean dose of 16.1 mg/kg/day for males and 15.7 mg/kg/day for females. There were no other findings considered to be of toxicological importance. A dietary level of 150 ppm (4.8 mg/kg/day in males; 4.6 mg/kg/day in females) was without toxicologically significant effects in male and female dogs.
Executive summary:

Groups of 4 male and 4 female beagle dogs were fed diets containing 0, 50, 150, or 500 ppm for a period of at least 1 year according to OECD guideline 452. Clinical observations and veterinary examination (including ophthalmoscopy) were made and body weights, food consumption and clinical pathology parameters were measured periodically throughout the study. At the end of the scheduled period, the animals were killed and subjected to a full examination post mortem. Selected organs were weighed and specified tissues were taken for subsequent histopathology examination. Dietary administration of 500 ppm to male and female dogs resulted in reduced growth and reduced food consumption. Administration of 500 ppm to female dogs resulted in an increased incidence of thin appearance. There was an increased incidence of reddened gums and of fluid faeces in males receiving 500 ppm but these were considered to be of no toxicological significance. At necropsy, group mean thyroid weights, adjusted for terminal body weight, for females receiving 500 ppm were higher than concurrent controls, however, in the absence of any associated treatment-related histopathological changes the increase in thyroid weight is considered to be of no toxicological significance. There were no changes in clinical pathology parameters and no pathology findings attributed to treatment with the test substance. 

Administration of 500 ppm in the diet for 52 weeks resulted in reduced growth and reduced food consumption in males and females. This dose was equivalent to an overall mean dose of 16.1 mg/kg/day for males and 15.7 mg/kg/day for females. There were no other findings considered to be of toxicological importance. A dietary level of 150 ppm (4.8 mg/kg/day in males; 4.6 mg/kg/day in females) was without toxicologically significant effects in male and female dogs.