Registration Dossier

Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
The purpose of this study was to assess the potential systemic toxicity in rats, including a screen for reproductive/developmental effects with administration of the target substance

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2013
Report Date:
2013

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: OECD Guideline 422
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder

Test animals

Species:
rat
Strain:
Wistar
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: SPF breeding, VELAZ
- Age at study initiation: 10 weeks on arrival
- Weight at study initiation: body weight - group mean ± standard deviation (g)
MALE
Group code 0 15 mg/kg bw 60 mg/kg bw
240 mg/kg bw
248.58 ± 10.33 246.23 ± 13.77 255.17 ± 11.91 243.32 ± 6.97
FEMALE
Group code 0 15 mg/kg bw 60 mg/kg bw
240 mg/kg bw
201.92 ± 6.38 200.73 ± 9.63 205.87 ± 5.15 199.53 ± 8.19
- Housing: 2 rats of the same sex in one cage in pre-mating period, during mating period – one male
and one female in one cage, pregnant females – individually, offspring – with mother, satellite
animals - 2 rats of the same sex in one cage.
- Bedding: sterilized soft wood fibers (Lignocel)
- Diet: complete pelleted diet for rats and mice in SPF breeding
- Water: drinking water ad libitum, quality corresponded to Regulation No. 252/2004 Czech Coll. of
Law, Health Ministry
- Acclimation period:6 days
- Health chech: During the acclimatisation period the health condition of all animals was controlled
daily. Then the animals were randomly divided into the control and test groups and they were ma
rked individually.
ENVIRONMENTAL CONDITIONS
- Temperature (°C):22 ± 3°C
- Humidity (%): 30-70%
- Air changes (per hr):
- Photoperiod: 12 hours cycle dark/light

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
Concentration Level 50 mg/10mL
Ca. 0.5 g of the test substance was weighted with wider end of glass Pasteur pipette into a 150mL g
lass beaker calibrated to 100 mL and the beaker was replenished by the vehicle. The solution was st
irred by magnetic stirrer (800 rpm) for 60 minutes.
Concentration Level 1000 mg/10 mL
Ca. 10 g of the test substance was weighted with wider end of glass Pasteur into a 150mL glass
beaker calibrated to 100 mL and the beaker was replenished by the vehicle and disslolved in ultr
asonic bath for a 15 min. This solution was stirred by magnetic stirrer (800 rpm) for 30 minutes.
Analytical verification of doses or concentrations:
no
Remarks:
not formulations were cheked during the in life part of the xperiment. However, homogeneity and stability of formulation analysis were performed. Parameters as linearity, reproducibiity, recovery are also evaluated
Details on analytical verification of doses or concentrations:
The stability and the homogeneity were determined by means of measuring of a peak area of the test substance by high-performance liquid chromatography based on the method developed at the test facility.
The stability of the application form The stability of the application form was checked by determination of a concentration of the test substance within 120 min (at the time 0 min, 30 min, 60 min and 120 min). The samples were taken from the middle of the beaker content after required time intervals.
The homogeneity of the application form The homogeneity of the application form was checked by determination of a concentration of the test substance in three places of solution (at the bottom, in the middle and at the surface).
Results of analysis
From the results of analyses (homogeneity and stability) follows that the solution of the test substance in vehicle in concentration level 50 mg/10 ml prepared at defined laboratory conditions (laboratory temperature, preparation of solution by defined manner) is homogenous and stable at least for 120 minutes starting with preparation of the application form. From the results of analyses (homogeneity and stability) follows that the solution of the test substance
in vehicle in concentration level 1000 mg/10 ml prepared at defined laboratory conditions (laboratory temperature, preparation of solution by defined manner) is homogenous and stable at least for 60 minutes from starting with analysis of the application form.
Duration of treatment / exposure:
Parental males:
1st day – 14th day (pre-mating) → 28th day (mating) → 42nd day of study
Satellite males:
329
1st day → 42nd day (administration) → 56th day (observation)
Parental females:
1st day – 14th day (pre-mating) → 28th day (mating) → gestation → lactation → day 4 post partum
Satellite females:
1st day → 42nd day (administration) → 56th day (observation)
Non-pregnant females (without evidence of copulation):
1st day – 14th day (pre-mating) → 28th day (mating) → 54th day of study
Non-pregnant females (with evidence of copulation):
1st day – 14th day (pre-mating) → 28th day (mating) → 25th day after confirmed mating (max. 54th day
of study)
Frequency of treatment:
7 days per week at the same time (8.00 – 10.00 am). The vehicle control group was administered by aqua pro injectione in the same volume.
Duration of test:
refer to duration of treatment
Doses / concentrationsopen allclose all
Dose / conc.:
15 mg/kg bw/day
Dose / conc.:
60 mg/kg bw/day
Dose / conc.:
240 mg/kg bw/day
No. of animals per sex per dose:
1. Control 0 12 males + 12 females
2. Low dose 15 mg/kg/day 12 males + 12 females
3. Intermediate dose 60 mg/kg/day 12 males + 12 females
4. High dose 240 mg/kg/day 12 males + 12 females
Satellite groups:
5. Control – vehicle – satellite: 0 6 males + 6 females
6. High dose – satellite: 240 mg/kg/day 6 males + 6 females
Control animals:
yes, concurrent vehicle

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: yes
males and females - daily during the administration period
All rats were observed daily during the administration period.
This observation was made in order to record possible clinical effects after application and all changes
in behaviour of animals. So it was done after application at the same time every day
(11.00 – 13.00 p.m.) – at the time of expectation of maximal effect of the test substance. Animals were
observed in natural conditions in their cages.
DETAILED CLINICAL OBSERVATIONS: Yes
before the first application and then weekly (except the mating period)
Functional observation: at the end of administration/observation period

BODY WEIGHT: Yes
males - weekly
females - weekly in premating and mating period,
during pregnancy: 0., 7th, 14th, 20th day, during lactation: 0. or 1st, 3rd and 4th day;

FOOD CONSUMPTION:
males - weekly (except the mating period)
females - weekly during premating period
during pregnancy and lactation – on the same days as body weight

WATER CONSUMPTION A: Yes
satellite males and females – twice a week

HAEMATOLOGY: Yes
at the end of administration/observation period
This examination was performed only in 6 males and 6 females of each group and in satellite males and females.

CLINICAL CHEMISTRY: Yes
at the end of administration/observation period
This examination was performed only in 6 males and 6 females of each group and in satellite males and females.

- Mortality control: twice daily
All rats during the treatment periods were examined for vitality or mortality changes twice daily.

GROSS PATHOLOGY: Yes
males and nonpregnant females – at the end of administration period

HISTOPATHOLOGY
Reproduction part of study (12 males and 12 females from each main group)
Pituitary gland
Ovaries
Uterus
Cervix of uterus
Vagina
Epididymis/Epididymides
Prostate gland
Seminal vesicles and coagulating gland
Testes
All gross lesions
Fetal examinations:
pups examination:
CAGE SIDE OBSERVATIONS: pups - as soon as possible after delivery and then daily
BODY WEIGHTS: pups (litters) – 0. or 1st, 3rd and 4th day;
GROSS PATHOLOGY :on the 4th day of lactation
Indices:
reproduction parameters – number of live born pups, number of corpora lutea, number of implantations, mean weight of pup on the 0./1st day and mean weight of pup on the 4th day

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
effects observed, treatment-related
Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Description (incidence):
No premature deaths occurred during the study.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
at the highest dose
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
There was no effect of treatment upon food intake.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
effects observed, treatment-related
Ophthalmological findings:
no effects observed
Haematological findings:
effects observed, non-treatment-related
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Description (incidence and severity):
only males examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
middle and high dose
Histopathological findings: neoplastic:
not examined
Other effects:
no effects observed
Description (incidence and severity):
There were no signs seen at the routine examination that were considered to be related to treatment with Amidoamine and no premature deaths occurred during the study.

Maternal developmental toxicity

Number of abortions:
effects observed, treatment-related
Description (incidence and severity):
middle and hgh dose levels
Pre- and post-implantation loss:
not examined
Total litter losses by resorption:
not examined
Early or late resorptions:
not examined
Dead fetuses:
not examined
Changes in pregnancy duration:
no effects observed
Description (incidence and severity):





Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): no effects observed
Field "Description (incidence and severity)" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.DescriptionIncidenceAndSeverityEffectsOnPregnancyDuration): All females allocated to study showed normal 4/5 day estrous cycles during the acclimatization period.
Estrous cyclicity, pre-coital interval, gestation length and index, mating performance and fertility were considered unaffected by treatment with Amidoamine (UVCB). All females showed diestrus at termination.
Changes in number of pregnant:
no effects observed
Description (incidence and severity):
All females allocated to study showed normal 4/5 day estrous cycles during the acclimatization period.
Estrous cyclicity, pre-coital interval, gestation length and index, mating performance and fertility were considered unaffected by treatment with Amidoamine (UVCB).
All females showed diestrus at termination
Other effects:
no effects observed
Details on maternal toxic effects:
All females of control group and the lowest and the middle dose level were mated. Evidence of copulation was not found in one female of the highest dose level.
The number of females achieving pregnancy was analogous in treated and control groups. Abortions occurred at the middle and the highest dose level. The duration of mating of females at the lowest and the middle dose level was similar to the control group. The duration of mating of females at the highest dose level was longer against control. The duration of pregnancy of females at all dose level was comparable to the control.
The number of females bearing live pups was well-balanced in the control and the lowest dose level. In females of the middle and the highest dose level this number was apparently decreased compared to control.
Mean number of corpora lutea was decreased at the highest dose level. Vice versa mean number of implantations was decreased at the middle dose level. Mean number of live pups at birth and at day 4 after parturition in treated mothers was similar to control.
Mean weight of the litter at the highest dose level was markedly decreased against control. Mean weights of pup recorded at the 1st check of litter after parturition and on 4th day of lactation was slightly decreased at the highest dose level.
Macroscopical findings were recorded in stomach of pups of treated mothers: sporadically at the lowest and the middle dose level and in more than 25% of pups at the highest dose level.
No significant differences of mating indexes were observed. Fertility indexes were decreased at the middle and the highest dose level. At the middle and the highest dose level gestation index was markedly decreased. Survival indexes of treated groups were analogous to control.
Pre-implantation losses were slightly increased at the middle dose level and post-implantation losses were increased at the middle and the highest dose level. No significant differences were detected in post-natal losses.



Effect levels (maternal animals)

Dose descriptor:
NOAEL
Effect level:
ca. 240 mg/kg bw/day (nominal)
Basis for effect level:
clinical signs
mortality
body weight and weight gain
food consumption and compound intake
water consumption and compound intake
haematology
clinical biochemistry
urinalysis
behaviour (functional findings)
organ weights and organ / body weight ratios
gross pathology
histopathology: non-neoplastic

Maternal abnormalities

Key result
Abnormalities:
no effects observed

Results (fetuses)

Fetal body weight changes:
not examined
Reduction in number of live offspring:
effects observed, treatment-related
Description (incidence and severity):
middle and high dose for number of bearing living pups
Changes in sex ratio:
not examined
Changes in litter size and weights:
effects observed, treatment-related
Changes in postnatal survival:
effects observed, treatment-related
External malformations:
no effects observed
Skeletal malformations:
no effects observed
Visceral malformations:
effects observed, treatment-related
Description (incidence and severity):
stomach at the highest dose

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
ca. 60 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
reduction in number of live offspring
changes in litter size and weights
changes in postnatal survival
external malformations
skeletal malformations
visceral malformations

Overall developmental toxicity

Key result
Developmental effects observed:
no

Applicant's summary and conclusion

Conclusions:
The substance was tested for developmental toxicity and the NOAEL is equal to 60 mg/kg/day