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Toxicological information

Repeated dose toxicity: oral

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Administrative data

Endpoint:
sub-chronic toxicity: oral
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
2008
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
GLP study according to OECD guideline 408 (adopted 1998). Minor restrictions: Ophthalmological examination not specifically stated in the document, but weekly investigation of eyes for mydriasis, miosis, and exophthalmos as well as histopathology of eyes after termination. Study was originally judged Klimisch 1. However, according to the "Practical guide 6: How to report read-across and categories" the maximum score for read-across is 2.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2009
Report Date:
2009

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
Deviations:
no
Qualifier:
according to
Guideline:
other: Commission Directive 96/54/EEC, B.26
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
-Name of test material (as cited in study report): Biofert Plusz
Details are presented in "Confidential details on test material"

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
-Source: Harlan Laboratories Ltd., CH
-Age at delivery: 7 weeks
-Weight at acclimatisation: 139 – 200 g
-Fasting period before study: No
-Housing: Groups of 5 animals in Macrolon type 4 cages
-Certified Diet ad libitum
-Tap water ad libitum
-Acclimation period: 7 days under test conditions after health examination. Only animals without any visible signs of illness were used

ENVIRONMENTAL CONDITIONS
-According to Guideline

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
The dose formulations were prepared weekly. Homogeneity of the test item in the vehicle was maintained during the daily administration period using a magnetic stirrer. The dose formulations were stored refrigerated in glass beakers (2 - 8°C).
Dose Volume: 10 mL/kg bw
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Results of the test item dose formulations analyses: range of 93.2 – 121.5 %. For 33 of 36 samples, the required content limit of +/- 20 % with reference to the nominal concentration was fulfilled (3 samples exceeded the upper range). All were found to be homogeneous, and all were found to be stable for four hours at room temperature and for seven days when refrigerated (2 - 8 °C).
Duration of treatment / exposure:
91 days
Frequency of treatment:
Once daily
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 100, 300 and 1000 mg/kg bw/d (dry mass). Daily dose levels for the liquid test product: 0, 356, 1068 and 3559 mg/kg bw/d
Basis:

No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
-Dose selection rationale: Based on a 14 d dose-range-finding study. High dose level corresponds to the limit dose recommended in OECD Guideline 408
-Rationale for animal assignment: Randomisation
Positive control:
no

Examinations

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

CLINICAL OBSERVATIONS
Yes: The animals were observed for clinical signs once before commencement of administration as well as twice daily on days 1 – 3 and once daily on days 4 – 91 (treatment period)

BODY WEIGHT
Yes: Body weights were recorded weekly during the acclimatization and treatment periods and before necropsy

FOOD CONSUMPTION AND COMPOUND INTAKE
Yes: The food consumption was recorded once during the acclimatization period and weekly thereafter

FOOD EFFICIENCY
Yes: The mean absolute food consumption and the mean relative food consumption of the treated rats compared with controls

OPHTHALMOSCOPIC EXAMINATION
No: However once weekly investigation of mydriasis, miosis, and exophthalmos

HAEMATOLOGY
Yes: Collection of blood from all animals (fasted) once after 13 weeks of exposure. Used anaesthetic was isoflurane. Examined parameters: According to guideline

CLINICAL CHEMISTRY
Yes: Time schedule for all animals (fasted): see haematology. Examined parameters: According to guideline

URINALYSIS
Yes: Urine was collected during the 18 hours fasting period (see haematology) into a specimen vial, using a metabolism cage. Examined parameters: According to guideline

DETAILED BEHAVIORAL OBSERVATIONS:
Yes: Once weekly (all rats): The animals were observed in their home cages, outside their home cages in a standard arena and in the hand. These observations were performed once before commencement of administration and once weekly (weeks 1 to 12) thereafter. Examined parameters: piloerection, salivation, hunched posture, ataxia, tremor/twitching, prostration, circling, spasm, hyperactivity, somnolence, increased exploration, reduced grooming, vocalisation, dyspnoea, tachypnoea, bradypnoea. Reflexes: blink, pinna, iridic light reflex, push-off (hind leg), pain response, startle/hearing, righting reflex. Other: lacrimation, limbs cyanotic, mydriasis, miosis, exophthalmos, reduced muscle tone

FUNCTIONAL OBSERVATION BATTERY
Yes: During week 13 (all animals). Relevant parameters: Grip strength and locomotor activity
Sacrifice and pathology:
GROSS PATHOLOGY
Yes: Organ wet weights as recommended in guideline

HISTOPATHOLOGY
Yes: As recommended in guideline
Other examinations:
no
Statistics:
Dunnett-test, Steel-test, Fisher's exact-test. Statistical analysis of: Body weights, clinical laboratory data, locomotor activity, grip strength, organ weights and ratios as well as macroscopic findings.

Results and discussion

Results of examinations

Clinical signs:
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:
no effects observed
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
Even at the highest dose tested (1000 mg/kg bw) no significant adverse effects were found. There were some minor changes in investigated parameters such as haematology or clinical chemistry, however these findings were considered to be within the range of normal biological variation and therefore without toxicological relevance.

Effect levels

Dose descriptor:
NOAEL
Effect level:
> 1 000 mg/kg bw/day (nominal)
Based on:
other: dry mass of the liquid test item
Sex:
male/female
Basis for effect level:
other: No adverse effects at the highest dose level tested (corresponds to the limit dose recommended in OECD Guideline 408)

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
In a 13 week gavage study according to OECD Guideline 408 in male and female Wistar rats, no effects of toxicological relevance were detected even at the high dose level of 1000 mg/kg bw/d (related to dry mass).