Registration Dossier

Toxicological information

Toxicity to reproduction

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Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Remarks:
based on test type (migrated information)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
07-11-2007 to 28-05-2008
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2008
Report Date:
2008

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
Qualifier:
according to
Guideline:
other: OPPTS 870.3650
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material: Lauramine oxide (Ammonyx LO)
- Physical state: Light to straw liquid
- Lot/batch No.: 7243678-T121
- Analytical purity: 30.27 %
- Stability under test conditions: In vehicle, 365 days
- Storage condition of test material: Room temperature (20 ± 5 ºC)

Test animals

Species:
rat
Strain:
other: HanRcc: WIST(SPF)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: RCC Ltd, Füllinsdorf, Switzerland.
- Age at study initiation: 11 weeks
- Weight at study initiation: Males, 294 to 330 g; females, 175 to 214 g.
- Housing: Individually in Makrolon type 3 cages with wire mesh tops and sterilised standard softwood bedding. During the pre-pairing period, cages with males were interspersed among those holding females to promote the development of regular estrus cycles.
- Diet: Pelletted standard mouse maintenance diet available ad libitum.
- Water: Community tap-water available ad libitum.
- Acclimation period: Under test conditions for 1 week after health examination.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3 ºC
- Humidity (%): 30 - 70 %
- Air changes (per hr): 10 - 15
- Photoperiod (hrs dark / hrs light): 12 h light/dark

IN-LIFE DATES: From: 07-11-2007 To: 03-01-2008

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
Test item was used as provided by the sponsor, adjusting the dosing formulations for its purity i.e. 30.27 %. The dosage formulations were prepared weekly using the test item as supplied by the Sponsor and using a correction factor of 3.3 due to the purity of 30.27 % of the test item. Lauramine oxide was weighed into a glass beaker on a tared precision balance and approximately 80 % of the vehicle was added (w/v). Using an appropriate homogeniser, a homogenous suspension was prepared. Having obtained a homogenous mixture, the remaining vehicle was added. Separate formulations were prepared for each concentration. Homogeneity of the test item was maintained during the daily administration period using a magnetic stirrer.
Details on mating procedure:
During the pairing period, females were housed with sexually mature males (1:1) in special automatic mating cages i.e. with synchronised timing to initiate the nightly mating period, until evidence of copulation was observed. This system reduced the variation in the copulation times of the different females. The females were removed and housed individually if the daily vaginal smear was positive for sperm, or a copulation plug was observed.
The day of mating was designated Day 0 post coitum.
Female that did not mate during the 14-day pairing period , was paired with a male of the same group which had already mated successfully.
All dams were allowed to give birth and rear their litters (F1 pups) up to Day 4 post partum. Day 0 post partum was designated as the day on which a female had delivered all her pups.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
On the first day of treatment samples from the control group as well as three samples (top, middle and bottom) of about 2 g of each concentration were taken prior to dosing for analysis of concentration and homogeneity. Samples of about 2 g of each concentration were taken from the middle only to confirm stability (4 h and 7 days). During the last week of the treatment, samples were taken from the middle to confirm concentration. The aliquots for analysis of dose formulations were frozen (-20 ± 5 ºC) and delivered on dry ice to be stored at -20 ± 5 ºC until analysis.
The samples were analysed by HPLC coupled to an ELSD detector following an analytical procedure developed at RCC and quantified with the area under the peak. The test item was used as the analytical standard. Analysed samples were not discarded without written consent from the study director.
The analytical part of the study was conducted at RCC Ltd, Itingen, Switzerland under GLP-compliant conditions. The identity of Lauramine oxide was confirmed by its retention time, which was similar to that measured in the working standards. The test item content in all samples was found to be in the accepted range of ± 20 % of the nominal content. In addition, the homogenous distribution of Lauramine oxide in highly purified water was demonstrated. The results of the analytical phase confirmed the correct preparation and storage of application formulations during the conduct of this study.
Duration of treatment / exposure:
Lauramine oxide was administered to male rats for at least 28 days and to female rats for 14 days prior to pairing, through the pairing and gestation period until the F1 generation reached Day 4 post partum.
Frequency of treatment:
Daily
Details on study schedule:
See table.
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 40, 100 and 250 mg/kg bw/day
Basis:
actual ingested
No. of animals per sex per dose:
10/sex/dose
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose levels were selected based on a previous dose range finding toxicity study in Han Wistar rats, RCC study No. B51592 in which dose levels of 30, 60, 120, 500 and 1000 mg/kg/day corrected for purity were tested. Both the 1000 and 500 mg/kg/day resulted in lethality after a single or two doses, respectively. The overall NOEL was 120 mg/kg/day (corrected for purity).
- Rationale for animal assignment: Computer-generated random algorithm was used, with body weights taken into consideration in order to ensure similar mean body weights in all groups.
10 mL/kg bw was administered.
Positive control:
No data

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily.
- Cage side observations included: Viability and mortality.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Once prior to the first administration of the test item and then weekly thereafter, detailed clinical observations were performed outside the home cage. Animals were observed for the following: Changes in skin fur, eyes, mucous membranes, occurrence of secretions and excretions and autonomic activity (e.g. lacrimation, ruffled fur, pupil size, unusual respiratory pattern). Changes in gait, posture and response to handling as well as the presence of clonic or tonic movements, stereotypes or bizarre behaviour were also reported. Additionally females were observed for signs of difficult or prolonged parturition and behavioural abnormalities during nesting and nursing.
At one time during the study (males shortly before the scheduled sacrifice and females on day 3 or 4 post partum) relevent parameters were performed with five P-generation males and five P-generation females randomly selected from each group. The Functional Observation Battery (FOB) assessment was conducted following the daily dosage administration.
Animals were observed for the following:
a) cage-side observations: Unusual body movements (e.g. tremours, convulsions) abnormal behaviour (e.g. circling, stereotypy) and posture as well as resistance to removal.
b) hand-held observations: Palpebral closure, pinna reflex, lacrimation, pupil size, pupil reactivity, salivation, muscle tone, extensor thrust response, righting reflex and reaction to handling.
c) Open field observations: Level of ambulatory activity, including rearing (one minute evaluation) responsiveness to sharp noise, paw pinch, gait evaluation, quantity of urine and faecal pellets voided.
d) Categorical observations (could be made at any time during the FOB): Hair coat, behaviour, respiration, muscle movements, eyes, hearing ability (Preyer's reflex) urine or faeces, soiling, general abnormalities, posture.
e) Measurements/counts: Hind limb/fore limb grip strength, landing food splay, rectal temperature.
Additionally, locomotor activity was measured quantitatively for the same animals. Activity was measured with an Activity Monitor AMS-0151. Activity of the animals (based on beam count) was recorded for 6 minute intervals over a period of 30 min. These data and the total activity over 30 min were reported.

BODY WEIGHT: Yes
- Time schedule for examinations: Recorded daily from treatment start to day of necropsy.

FOOD CONSUMPTION:
- Food consumption:
Males, weekly during pre-pairing periods and after pairing periods.
Females, prepairing period days 1-8 and 8-14; gestation days 0-7, 7-14 and 14-21 post coitum, and days 1-4 post partum. No food consumption was recorded during the pairing period.

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 AND COMPOUND INTAKE: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Blood samples were obtained on the day before or the day of the scheduled necropsy from 5 males (randomly selected) from each group. Blood samples for 5 lactating females (randomly selected) from each group were obtained on day 5 post partum. The samples were collected early in the working day to reduce biological variation caused by circadian rhythms.
- Anaesthetic used for blood collection: Yes, isoflurane.
- Animals fasted: Yes, approximately 18 h before blood sampling but allowed access to water ad libitum.
- How many animals: 5 males (randomly selected) from each group. Blood samples for 5 lactating females (randomly selected) from each group were obtained on day 5 post partum.
- Parameters examined:
Complete blood cell count: Erythrocyte count, haemoglobin, hematocrit, mean corpuscular volume, red cell volume distribution width, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, haemoglobin concentration distribution width, leukocyte count (total), differential leukocyte count, platelet count.
Coagulation: Thrombin time (= thromboplastin time), activated partial thromboplastin time.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Blood samples were obtained on the day before or the day of the scheduled necropsy and for lactating females (randomly selected) from each group obtained on day 5 post partum. The samples were collected early in the working day to reduce biological variation caused by circadian rhythms.
- Animals fasted: Yes, approximately 18 h before blood sampling but allowed access to water ad libitum.
- How many animals: 5 males (randomly selected) from each group. Blood samples for 5 lactating females (randomly selected) from each group were obtained on day 5 post partum.
- Parameters examined: Glucose, urea, creatinine, bilirubin (total) cholesterol (total) triglycerides, aspartate aminotrasferase, alanine aminotransferase, alkaline phosphatase, gamma-glutamyl transferase, bile acids, creatine kinase, sodium, potassium, chloride, calcium, phosphorous, protein (total) albumin, globulin, albumin/globulin ratio.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: See section above on clinical signs.
Oestrous cyclicity (parental animals):
During the pre-pairing period , cages with males were interspersed among those holding females to promote the development of regular estrous cycles.
Sperm parameters (parental animals):
The reproductive organs were given particular attention during necropsy. The testes and epididymides of all parental males were weighed as pairs. The prostate, seminal vesicles with coagulating gland, testes and epididymides were preserved from all parental males. The testes were examined by PAS-hematoxylin. Histopathological examination placed special emphasis on stages of spermatogenesis and histopathology of interstitial cell structure.
Litter observations:
PARAMETERS EXAMINED
The following parameters were examined in F1 offspring: Litter size, live births and any gross anomalies, sex ratio, individual pup weights on Day 0 (when possible) and on Day 1 and Day 4 post partum.

GROSS EXAMINATION OF DEAD PUPS: Yes except those excessively cannibalised.
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All animals sacrificed after treatment of at least 28 days, when no longer needed for assessment of reproductive effects or found dead.
- Maternal animals: All animals sacrificed on day 5 post partum or found dead.

GROSS PATHOLOGY: Specimens of abnormal tissue were fixed in neutral phosphate buffered 4 % formaldehyde solution.
For the parental animals, special attention was directed to the organs of the reproductive system. The number of implantation sites and corpora lutea were recorded for all dams with litters. The uteri of non-pregnant females were placed in a solution of ammonium sulfide to visualize possible haemorrhagic areas of implantation sites.

Organ weights.
The testes and epididymides of all parental males were weighed as pairs. In addition, from 5 males and females selected randomly from each group, the following organs were trimmed from any adherent tissue, as appropriate, and their wet weight recorded.
Adrenal glands (weighed as pairs), brain, heart, kidneys (weighed as pairs), liver, thymus and spleen.
The following tissues from all parental males were preserved in neutral phosphate buffered 4 % formaldehyde solution or in Bouin's fixative:
Prostate, seminal vesicles with coagulating gland, testes (in Bouin's fixative) and epididymides (in Bouin's fixative).
Ovaries from all parental females were preserved in neutral phosphate buffered 4 % formaldehyde solution.
In addition, from 5 males and 5 females per group selected for organ weights, the following tissues were preserved in neutral phosphate buffered 4 % formaldehyde solution: Gross lesions, brain, spinal chord, small and large intestine (including Peyer's patches), stomach, liver, kidneys, adrenals, spleen, heart, thymus, thyroids and parathyroids (if possible) trachaea and lungs (preserved by inflation with fixative and then immersion) uterus with vagina, urinary bladder, lymph nodes (mesenterial, mandibular) peripheral nerve (sciatic) and bone marrow.


HISTOPATHOLOGY: Yes all organ and tissue samples were processed, embedded and cut at an approximate thickness of 2 - 4 μm and stained with hematoxylin and eosin. Additionally, the testes were examined by PAS-hematoxylin.
Slides of all organs and tissues listed above and collected at terminal sacrifice from the animals of the control and high-dose groups were examined by the study pathologist. The same applied to all occurring gross lesions and to all animals which died spontaneously.
Special emphasis was placed on stages of spermatogenesis and histopathology of interstitial cell structure.
If test item-related morphological changes were detected in organs of any high-dose animal, those same organs from the mid- and low-dose group were examined to establish a no-effect level, if possible.
Histological examination of ovaries was carried out on any females that did not give birth.
Postmortem examinations (offspring):
SACRIFICE: Day 4 post partum.

GROSS NECROPSY: Dead pups, except those excessively cannibalised, were examined macroscopically.
Statistics:
The following statistical methods were used for food consumption, body weight, macroscopical findings, organ weights and reproduction data, locomotor activity, rectal temperature, landing foot splay, grip strength, haematology and clinical chemistry:
Mean and standard deviations of various data were calculated and included in the report.
The Dunnett-test (many to one t-test) based on pooled variance estimate was applied if the variables could be assumed to follow a normal distribution for the comparison of the treated groups and the control groups for each sex.
The Steel-test (many-one rank test ) was applied instead of the Dunnett-test when the data could not be assumed to follow a normal distribution.
Fisher's exact test was applied if the variables could be dichotomized without loss of information.
Reproductive indices:
The following reproduction parameters were calculated: Fertility indices, mean precoital time, post-implantation losses, mean litter size, pup sex ratios and viability indices.
For reproduction data, group mean values were calculated both on a litter basis and on a percentage per group basis. Mean pup weights were calculated from individual weights both on a per group and on a per litter basis.
Offspring viability indices:
The following offspring viability indices were calculated: Mean litter size, pup sex ratios and viability indices. Mean pup weights were calculated from individual weights both on a per group and on a per litter basis.

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
no effects observed
Other effects:
not specified

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
no effects observed
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
effects observed, treatment-related

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
At 250 mg/kg/day, one female was found dead at the beginning of the gestation period. This was not considered to be a test item-related effect, since no adverse clinical signs were noted before and it was a single case. At 100 and 250 mg/kg/day all males were noted to have reduced activity. Rales and salivation were noted occassionally at 250 mg/kg/day. In females, rales and salivation were noted in three females at 250 mg/kg/day during the gestation period.
Functional Observational Battery:
At 250 mg/kg/day, total locomotor activity was statistically significantly reduced in females.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
Males at 250 mg/kg/day mean body weight and mean body weight gain were reduced throughout the study. At 100 mg/kg/day a decrease was noted during the pre-pairing period. In females at 250 mg/kg/day mean body weight and mean body weight gain were generally decreased for the whole study.
Males at 250 mg/kg/day mean food consumption was dose-dependently reduced throughout the study treatment period. In females, at 100 and 250 mg/kg/day, mean food consumption was dose-dependently reduced during the pre-pairing period and gestation period. During the lactation period, at 100 mg/kg/day it recovered and at 250 mg/kg/day remained reduced.

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
No data

REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS)
No data

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
All pairs mated. Mean pre-coital time, fertility index and conception rates were not affected by the treatment with the test item. At 250 mg/kg/day gestation index was reduced since two dams did not deliver any pups and one dam delivered only one dead pup.
Implantation rate was unaffected. At 250 mg/kg/day, a statistically significant increase of post-implantation loss was observed.

ORGAN WEIGHTS (PARENTAL ANIMALS)
At 250 mg/kg/day, an increase in the absolute and relative liver weight was noted in males and females.

GROSS PATHOLOGY (PARENTAL ANIMALS)
At 250 mg/kg/day, at necropsy, the mucosa in the forestomach was thickened and with irregular surface and a thickened stomach was observed in 5 of 10 males.

HISTOPATHOLOGY (PARENTAL ANIMALS)
Microscopically the test item-related lesions recorded were:
Spleen: Lymphoid depletion was noted in females (2/5) at 250 mg/kg/day.
Liver: Hepatocellular hypertrophy in males (2/5) and females (1/5) at 250 mg/kg/day.
Kidneys: An increase of tabular basophilia and hyaline droplets was recorded in treated males. However, this increase in incidence was considered below the scope of concern in males receiving 40 or 100 mg/kg/day since they showed the same severity grade as control males. An increase in severity grade of both findings was only recorded in males treated at 250 mg/kg/day.
Forestomach: Hyperkeratosis, parakeratosis, squamous cell hyperplasia and submucosal inflammation were recorded in all males and females at 100 and 250 mg/kg/day.
Submucosal oedema was recorded in males (3/5) and in females (3/5) at 250 mg/kg/day and in females at 100 mg/kg/day.
Erosions were recorded in males (3/5) and females (3/5) at 250 mg/kg/day.
Ulcerations were recorded in females at 100 (1/5) and 250 mg/kg/day (1/5).
Pustules were recorded in males at 250 mg/kg/day (2/5) and females at 100 (1/5) and 250 mg/kg/day.
Microscopic examination of the reproductive organs of males and females treated at 250 mg/kg/day failed to find any abnormality.

OTHER FINDINGS (PARENTAL ANIMALS):
CLINICAL LABORATORY INVESTIGATIONS: The statistically significant alterations observed at 250 mg/kg/day were not considered to be adverse.

Effect levels (P0)

open allclose all
Dose descriptor:
NOAEL
Effect level:
40 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Dose descriptor:
NOEL
Effect level:
100 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Based on reproduction/developmental toxicity.

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings:
no effects observed

Details on results (F1)

VIABILITY (OFFSPRING)
Litter size, sex ratio and abnormalities at first litter check, postnatal loss Day 0 - 4 post partum: Litter size and mean number of pups at first litter check were not affected by the treatment with the test item. At 250 mg/kg/day, a statistically significant increase in pup death was observed on postnatal days 0 - 4. Although the higher postnatal loss was in the range of historical control data this resulted in a reduced number of pups.

CLINICAL SIGNS (OFFSPRING)
No abnormal pup was noted at any dose level, except at 250 mg/kg/day where mean pup weight development was reduced.

GROSS PATHOLOGY (OFFSPRING)
At necropsy of pups, there were no abnormal findings.

HISTOPATHOLOGY (OFFSPRING)
At necropsy of pups, there were no abnormal findings.

OTHER FINDINGS (OFFSPRING)
The sex ratio was also not affected.

Overall reproductive toxicity

Reproductive effects observed:
not specified

Any other information on results incl. tables

Summary of results:

Parameter

Administration dose

Control group

Low dose group

Medium

dose group

High

dose group

mg/kg

0

40

100

250

Sex

M

F

M

F

M

F

M

F

No. of animals

10

10

10

10

10

10

10

10

Mortality

0

0

0

0

0

0

1*

0

Tolerability

Reduced activity

-

-

-

-

+

-

+

-

Rales and salivation

-

-

-

-

-

-

+

+

(gestation period)

FOB

Total locomotor activity

-

-

-

-

-

-

-

+

Food consumption

Reduction

-

-

-

-

+

+

(pre-pairing & gestation)

+

+

(pre-pairing & gestation)

Recovery

-

-

-

-

-

+

-

-

Body weight

-

-

-

-

-

-

+

+

Body weight gain

-

-

-

-

-

-

+

+

Clinical Laboratory Investigation

-

-

-

-

-

-

-

-

Reproduction data

Mean pre-coital time

-

-

-

-

Implantation rate

-

-

-

-

Conception rate

-

-

-

-

Gestation index

-

-

-

+

Organ weights

Liver weight increase

Absolute

-

-

-

-

-

-

+

+

Liver weight increase

Relative

-

-

-

-

-

-

+

+

Macroscopic findings and histopathology

Thickened mucosa in the forestomach (n)

-

-

-

-

-

-

5

-

Microscopic lesions

Spleen: Lymphoid depletion (n)

-

-

-

-

-

-

-

2

Liver: Hepatocellular hypertrophy (n)

-

-

-

-

-

-

2

1

Kidney: Increased incidence of tubular basophilia and hyaline droplets

-

-

+ ns

-

+ ns

-

+

-

Forestomach: Hyperkeratosis

-

-

-

-

+

+

+

+

Forestomach: Parakeratosis

-

-

-

-

+

+

+

+

Forestomach: Squamous cell hyperplasia

-

-

-

-

+

+

+

+

Forestomach: Submucosal inflammation

-

-

-

-

+

+

+

+

Forestomach: Submucosal edema (n)

-

-

-

-

-

2

3

3

Forestomach: Erosions (n)

-

-

-

-

-

-

3

3

Forestomach: Ulcerations (n)

-

-

-

-

-

1

-

1

Forestomach: Pustules (n)

-

-

-

-

-

1

2

1

Microscopic Reproductive abnormalities

-

-

-

-

-

-

-

-

Litter data

Litter size

-

-

-

-

Mean no. of pups at first litter check

-

-

-

-

Sex ratio

-

-

-

-

Pup abnormalities

-

-

-

-

Pup death postnatal Day 4

-

-

-

+

Decreased Pup weight Day 4

-

-

-

+ ns

Pup necropsy findings

-

-

-

-

*Not considered to be a test item-related effect.

ns: not significant

Applicant's summary and conclusion

Conclusions:
The Lauramine oxide was administered in highly purified water as vehicle, at dosages of 40, 100 and 250 mg/kg/day, corrected to 30.27 % purity and controls received the vehicle only over a number of consecutive weeks. Lauramine oxide was administered to male rats for at least 28 days and to female rats for 14 days prior to pairing, through pairing and gestation period until the F1 generation reached Day 4 post partum.
Administration at 100 and 250 mg/kg bw /day caused a reduction in activity and of the body weight gain in males during the pre-pairing period and a dose-dependent reduction of food consumption in males and females. At 250 mg/kg bw/day treatment with the test item resulted in a general reduction of body weight gain in males and females, statistically reduced locomotor activity in females and in statistically significantly uncreased post-implantation and postnatal loss and in decreased pup body weight development. An increase of liver weights (absolute and ratios) was observed and correlated with hepatocellular hypertrophy noted during the histopathological examination. At necropsy, the mucosa in the forestomach was thickened with an irregular surface. The histopathological data showed lesions in the forstomach and in the kidneys. At 100 mg/kg bw/day, lesions in the forestomach were noted at necropsy and histopathology. Based on these results the overall NOAEL general was established at 40 mg/kg/day. The NOEL for reproduction/developmental toxicity was considered to be 100 mg/kg/day.