Registration Dossier

Toxicological information

Repeated dose toxicity: oral

Currently viewing:

Administrative data

Endpoint:
chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
before May 1977
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
(no analytical verification of concentrations)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1977
Report Date:
1977

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 452 (Chronic Toxicity Studies)
Version / remarks:
The study was conducted at a time before the specified guideline was available. However, the design is similar to the OECD TG 452.
Principles of method if other than guideline:
Chronic feeding study, 20 male and 20 female rats/group were fed diets for 52 weeks, additional 10 male and 10 female sentinel animals were killed after 26 weeks for interim pathological examination, diets with 0.08, 0.4, and 2 % test material (corresponding to daily doses of approx. 40, 200, and 1000 mg/kg bw), endpoints investigated: clinical signs, body weights, compound intake, ophthalmoscopy, haematology, clinical chemistry, urinalysis, gross necropsy, organ weights, histology.
GLP compliance:
no
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid
Specific details on test material used for the study:
Information from study report:
- Name of test material: Hostapur (SAS 60)
- Appearance/Further information: aqueous slurry (60 % w/w active ingredient); Hostapur (SAS 60) is equal to 60 % Hostapur SAS 93 in water
- Batch number: batch received on 3 September 1974 (lot number NR T 2/112)
- Purity (of Hostapur SAS 93): 91-95 % active ingredient
- Solubility and stability of the test substance in the diet: No data

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on species / strain selection:
Sprague Dawley CD strain
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Strain: Sprague Dawley CD strain
- Source: Charles River U.K. Limited, Margate, England
- Weight at study initiation (mean): males 111-119 g, females 109-110 g
- Housing: 5 animals per cage, polypropylene cages with stainless steel mesh floors
- Diet and water: ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 2
No further data

Administration / exposure

Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Remarks:
substance given in complete rodent diet
Details on oral exposure:
All concentrations in the report were quoted in terms of the active ingredient. For converting substance concentrations in feed into daily doses the factor of 0.05 was used (default factor for chronic studies for rats; Guidance on selected default values to be used by the EFSA Scientific Committee, Scientific Panels and Units in the absence of actual measured data, EFSA Journal 2012;10(3):2579)

DIET PREPARATION
- Rate and description of preparation of diet: The material was heated to 50 °C, and agitated for 10 hours to achieve homogeneity, before removing samples. Pre-mixes with powdered diet were prepared weekly and dried overnight at 30 °C; the requisite quantities of this pre-mix were then diluted with fresh powdered diet and mixed in a rotary double-cone mixer for 20 minutes to achieve homogeneity.
- Type of food: complete rodent diet (Spratts Laboratory diet no. 2)
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
52 weeks at maximum; animals of satellite groups were treated for 26 weeks and then killed for interim pathological examination.
Frequency of treatment:
not applicable, diet was freely available to rats
Doses / concentrationsopen allclose all
Dose / conc.:
40 mg/kg bw/day (nominal)
Remarks:
Corresponding dose for group receiving diet with 0.08 % (w/w) test substance
Dose / conc.:
200 mg/kg bw/day (nominal)
Remarks:
Corresponding dose for group receiving diet with 0.4 % (w/w) test substance
Dose / conc.:
1 000 mg/kg bw/day (nominal)
Remarks:
Corresponding dose for group receiving diet with 2.0 % (w/w) test substance
No. of animals per sex per dose:
20; additional 10 per sex and dose were used as satellite animals (for interim pathological examination)
Control animals:
yes, plain diet
Details on study design:
Groups of 30 male and 30 female rats were fed diets for 52 weeks at maximum containing 0, 0.08, 0.4, and 2.0 % (w/w) substance. For each dose 10 male and 10 female sentinel animals were killed after 26 weeks of treatment for interim pathological examination.

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
All rats were observed daily for ill health, and examined for any evidence of reaction to treatment in respect of behaviour, appearance and locomotion.

BODY WEIGHT: Yes
- Time schedule for examinations: on Day 0, at weekly intervals for the first 13 weeks, and at monthly intervals thereafter.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
The quantity of food eaten by each cage of rats was calculated weekly, by measurement of the food residues.

OPHTHALMOSCOPIC EXAMINATION: Yes
Both eyes of each rat were examined, using a Keeler direct ophthalmoscope, prior to the commencement of treatment. After 26 and 52 weeks of treatment, all rats due for sacrifice were again examined.

HAEMATOLOGY: Yes
After 26 and 52 weeks of treatment, blood samples were taken from ten male and ten female rats from each group for evaluation of the following characteristics: Packed cell volume (PCV), Haemoglobin concentration (Hb), Erythrocyte count (RBC), Leucocyte count (WBC) - total, Leucocyte count (HBC) - differential, Mean cell haemoglobin concentration (MCHC), Mean cell volume (MCV).

CLINICAL CHEMISTRY: Yes
After 26 and 52 weeks of treatment, blood samples were taken from five males and five females from each group for evaluation of the following parameters: Urea concentration, Glucose concentration, Total protein concentration, Electrophoretic protein fractions, Alkaline phosphatase activity (SAP), Glutamate-pyruvate transaminase activity (SGPT), Sodium (Na) and potassium (K) concentrations.

URINALYSIS: Yes
After 26 and 52 weeks of treatment five male and five female rats from each group were placed in individual metabolism cages for collection of urine samples. Drinking water was removed at 1500 hours and urine collected from 1700 hours to 0900 hours the following day; the concentrated sample was centrifuged at 3,400 r.p.m. for eight minutes.
- Parameters checked in supernatant: pH, Specific gravity (SG), Protein concentration, Reducing substances, Glucose, Ketones, Blood pigments, Bile pigments, Urobilinogen.
- Parameters checked in deposit: Epithelial cells (E), Polymorphonuclear leucocytes (P), Mononuclear leucocytes (N), Erythrocytes (R), Casts (C), Other abnormal constituents (A).

OTHER: An Urine concentration test was conducted. After 26 and 52 weeks of treatment the volume and specific gravity of urine samples collected over a two-hour period after water deprivation for six hours (0800 - 1400 hours) were examined.
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
Each rat found dead or sacrificed during the test was subjected to a detailed macroscopic examination post mortem.
The weights of the following organs were recorded for all rats: Adrenals, Brain, Gonads, Heart, Kidneys, Liver, Lungs, Pituitary, Spleen, Thyroids.

HISTOPATHOLOGY: Yes
A full spectrum of tissues was preserved where possible, and macroscopically abnormal tissues were examined histologically in an attempt to define the cause of death.
Both glandular and non-glandular areas of the stomach, auricular and ventricular sections of the heart and three levels of the brain (cerebellum, cerebral cortex and medulla) were prepared in section.
Microscopic examination of all tissues from all animals was carried out.
Statistics:
no data

Results and discussion

Results of examinations

Details on results:
CLINICAL SIGNS AND MORTALITY
Eight rats died during the test period; the cause of death or incidence of mortality in the groups were unrelated to treatment with the test substance.
All rats given diets containing 2.0 % w/w (approx. 1000 mg/kg) displayed a lack of grooming activity. By the end of the fourth week, the leaner body conformation of rats at the high dose level was discernible on handling.
Rats of either sex given the intermediate or low treatment levels were indistinguishable from controls.

BODY WEIGHT AND WEIGHT GAIN
Decreased rates of bodyweight gain were seen in males and females given diets containing 2.0% w/w (approx. 1000 mg/kg) throughout the treatment period, but not in the lower dose groups.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
Food intake was reduced in males at the highest level over the first 19 weeks. Females at the highest level and rats of both sexes at lower levels were unaffected in this respect.

OPHTHALMOSCOPIC EXAMINATION
No substance-related effects reported.

HAEMATOLOGY
not affected by the treatment

CLINICAL CHEMISTRY
Serum alkaline phosphatase (SAP) and serum glutamate-pyruvate transaminase (SGPT) was marginally increased in animals receiving the highest dietary concentration at week 26 and 52. In the absence of structural hepatic or other changes (vide infra) that might have accounted for such minor disturbances, it was concluded that the increased activities were not of toxicological significance.

URINALYSIS
not affected by the treatment

ORGAN WEIGHTS
No disturbances of absolute and relative organ weights relating to treatment were seen in rats killed after 26 or 52 weeks of treatment.

GROSS PATHOLOGY AND HISTOPATHOLOGY: NON-NEOPLASTIC
Necropsy findings of rats killed after 26 or 52 weeks revealed no changes in organ morphology attributable to treatment. No significant histopathological change or variation from normal was seen in the tissues examined that was considered to be related to the administration of the test compound. Minor changes in the liver and kidney were considered to reflect the changed nutritional status of animals receiving the highest dose level.

OTHERS: Urine concentrating ability was not affected by the treatment.

Effect levels

Key result
Dose descriptor:
NOAEL
Effect level:
ca. 200 mg/kg bw/day (nominal)
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
clinical signs
body weight and weight gain

Target system / organ toxicity

Critical effects observed:
yes
Lowest effective dose / conc.:
1 000 mg/kg bw/day (nominal)
System:
other: impaired grooming activity and retarded body weight gain
Organ:
other: impaired grooming activity and retarded body weight gain
Treatment related:
yes

Applicant's summary and conclusion

Executive summary:

A chronic feeding study similar to OECD TG 452, but conducted at a time before the guideline was available, investigated the chronic toxicity of the substance (test material was as aqueous slurry with 60 % active ingredient; all concentrations quoted here are in terms of the active ingredient). For that purpose groups of 20 male and 20 female rats were fed diets containing 0.08, 0.4 and 2.0 % (w/w) test substance for 52 weeks (approximate doses 40, 200 and 1000 mg/kg bw and day). A similar sized group received plain diet and served as control. For each dose additional 10 male and 10 female sentinel animals were used for interim pathological examination (terminated after 26 weeks of treatment).

As result it was demonstrated, that concentrations of the test substance in the diet up to 0.4 % (w/w; approx. dose 200 mg/kg bw) were tolerated by the animals without any significant effect. Even at the highest dose level of 2 % (w/w; approx. 1000 mg/kg) only unspecific effects not accompanied by any functional or structural changes have been observed (i.e. impaired grooming activity and retarded body weight gain). No NOAEL was stated in the report, but based on the available data a NOAEL of 200 mg/kg bw (corresponding to diet with 0.4 %) can be defined.