Registration Dossier

Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From October 11 to November 25, 2011
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
Study performed according to OECD test guideline No. 421 and in compliance with GLP.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report Date:
2012

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
Deviations:
yes
Remarks:
High dose level was reduced from 1000 mg/kg bw/day to 500 mg/kg bw/day on Day 3 due to toxicity
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes (incl. certificate)
Remarks:
UK GLP Compliance Programme (inspected on July 19-21, 2011/ signed on August 31, 2011)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Details on test material:
- Physical state: Clear colourless liquid
- Storage condition of test material: Room temperature in the dark, under nitrogen

Test animals

Species:
rat
Strain:
Wistar
Remarks:
Wister HanTM: RccHanTM: WIST strain
Details on species / strain selection:
The rat was selected for this study as it is a readily available rodent species historically used in safety evaluation studies and is acceptable to appropriate regulatory authorities.
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Laboratories U.K. Ltd., Blackthorn, Bicester, Oxon, UK.
- Age at study initiation: Approximately 12 weeks
- Weight at study initiation: (P) Males: 326-360 g; Females: 189-221 g
- Housing: Animals were housed in groups of five in solid floor polypropylene cages with stainless steel mesh lids and softwood flake bedding (Datesand Ltd., Cheshire, UK). During the mating phase, the animals were transferred to polypropylene grid floor cages suspended over trays lined with absorbent paper on a 1 male: 1 female basis. Following evidence of successful mating, the males were returned to their original cages. Mated females were housed individually during gestation and lactation, in solid floor polypropylene cages with stainless steel mesh lids and softwood flakes.
- Diet (e.g. ad libitum): A pelleted diet (Rodent 2018C Teklad Global Certified Diet, Harlan Laboratories U.K. Ltd., Oxon, UK), ad libitum
- Water (e.g. ad libitum): Mains drinking water, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 2 °C
- Humidity (%): 55 ± 15%
- Air changes (per hr): At least fifteen air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hrs dark / 12 hrs light

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
arachis oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Test item was prepared at the appropriate concentrations as a solution in Arachis oil BP. Formulations were prepared based on available stability data either daily, weekly or fortnightly and stored at approximately 4 ºC in the dark under nitrogen.

VEHICLE
- Concentration in vehicle: 0, 15, 62.5, 250*/125 mg/mL (* Following the early termination of three high dose level animals on Day 3 of the study the high dose level was reduced from 1000 mg/kg bw/day to 500 mg/kg bw/day. Treatment was discontinued for the high dose animals on Day 4 of the study and recommenced on Day 5 at the adjusted dose level. As for the majority of the duration of this study high dose level of 500 mg/kg bw/day was used it is reported as such.)
- Amount of vehicle (if gavage): 4 mL/kg bw/day
Details on mating procedure:
- M/F ratio per cage: 1 male: 1 female basis
- Length of cohabitation: For a maximum of 14 days
- Proof of pregnancy: Vaginal plug and sperm in vaginal smear referred to as day 0 of pregnancy.
- After successful mating each pregnant female was caged (how): Mated females were housed individually during the period of gestation and lactation.

Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples of each test item formulation were taken and analysed for concentration of test item. The results indicate that the prepared formulations were within ± 2% of the nominal concentration.
Duration of treatment / exposure:
Up to 8 weeks (including a two week maturation phase, pairing, gestation, and early lactation for females)
Male: Days 1-43 (mating on Days 15/16)
Female: Days 1 to post partum Day 5 (mating on Days 15/16)

After the initial decline of animal health, the high dose level was reduced from 1000 mg/kg bw/day to 500 mg/kg bw/day following agreement with the Sponsor. Animals in the high dose group were not dosed on Day 4 of the study, and re-started dosing on Day 5 at the lower dose of 500 mg/kg bw/day. Afterwards, animals at 500 mg/kg bw/day level showed recovery of the previously observed clinical signs.
Frequency of treatment:
Daily
Details on study schedule:
Chronological Sequence of Study:

i) Groups of ten male and ten female animals were treated daily at the appropriate dose level throughout the study (except for 1000 mg/kg bw/day animals which were not dosed on Day 4 prior to a dose reduction to 500 mg/kg bw/day and females during parturition where applicable). The first day of dosing was designated as Day 1 of the study.

ii) On Day 15 (Treatment Groups: Control, Low and Intermediate) or Day 16 of the study (Treatment Group: High), animals were paired on a 1 male: 1 female basis within each dose group for a maximum of fourteen days.

iii) Following evidence of mating (designated as Day 0 post coitum) the males were returned to their original cages and females were transferred to individual cages.

iv) Pregnant females were allowed to give birth and maintain their offspring until Day 5 post partum. Evaluation of each litter size, litter weight, mean offspring weight by sex, clinical observations and landmark developmental signs were also performed during this period.

v) The male dose groups were killed and examined macroscopically on Day 43 of the study.

vi) At Day 5 post partum, all surviving females and surviving offspring were killed and examined macroscopically.
Doses / concentrationsopen allclose all
Dose / conc.:
60 mg/kg bw/day (actual dose received)
Dose / conc.:
250 mg/kg bw/day (actual dose received)
Dose / conc.:
500 mg/kg bw/day (actual dose received)
Remarks:
initially 1000 mg/kg bw/day, reduced to 500 mg/kg bw/day on Day 5
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose levels were chosen based on the results of previous toxicity work, i.e., the oral 28-day repeat dose toxicity study in Sprague-Dawley rats (Study Number FIR/062).
- Rationale for animal assignment (if not random): The animals were allocated to dose groups using a randomisation procedure based on stratified body weights and the group mean body weights were then determined to ensure similarity between the dose groups.
Positive control:
None.

Examinations

Parental animals: Observations and examinations:
CLINICAL OBSERVATIONS: Yes
- Time schedule: All animals were examined for overt signs of toxicity, ill-health and behavioural change immediately before dosing, soon after dosing, and one and five hours after dosing, during the working week (one hour after dosing at weekends (except for females during parturition where applicable).

BODY WEIGHT: Yes
- Time schedule for examinations: Individual body weights were recorded on Day 1 (prior to dosing) and then weekly for males until termination and weekly for females until mating was evident. Body weights were then recorded for females on Days 0, 7, 14 and 20 post coitum, and on Days 1 and 4 post partum.

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes.
- During the pre-mating period, weekly food consumption was recorded for each cage of adults until pairing. This was continued for males after the mating phase. For females showing evidence of mating, food consumption was recorded for the periods covering post coitum Days 0-7, 7-14 and 14-20. For females with live litters, food consumption was recorded during the lactation period (Days 1-4).
Weekly food efficiency (body weight gain/food intake) was calculated retrospectively for males and for females during the pre-mating phase. Due to offspring growth and milk production, food efficiency could not be accurately calculated during gestation and lactation.

WATER CONSUMPTION: Yes
- Time schedule for examinations: Water intake was observed daily by visual inspection of water bottles for any overt changes.

OTHERS:
Pregnancy and Parturition: Each pregnant female was observed at approximately 0830, 1230 and 1630 hours and around the period of expected parturition. Observations were carried out at approximately 0830 and 1230 hours at weekends and public holidays.
Oestrous cyclicity (parental animals):
Not examined
Sperm parameters (parental animals):
Not examined
Litter observations:
PARAMETERS EXAMINED
The following parameters were examined in F1 offspring: Number of offspring born (both live and stillborn), Number of offspring alive recorded daily and reported on Days 1 and 4 post partum, Sex of offspring on Days 1 and 4 post partum, Clinical condition of offspring from birth to Day 5 post partum, Individual offspring weights on Days 1 and 4 post partum (litter weights were calculated retrospectively from this data)
All live offspring were assessed for surface righting reflex on Day 1 post partum.

GROSS EXAMINATION OF DEAD PUPS: yes, for external and internal abnormalities; possible cause of death was not determined for pups born or found dead.
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals were killed by intravenous overdose of sodium pentobarbitone followed by exsanguination on Day 43.
- Maternal animals: Adult females were killed by intravenous overdose of sodium pentobarbitone followed by exsanguination on Day 5 post partum. Any females which failed to achieve pregnancy or produce a litter were killed on or after Day 26 post coitum.

GROSS NECROPSY
- Gross necropsy consisted of a full external and internal examination. For all females, the uterus was examined for signs of implantation and the number of uterine implantations in each horn was recorded.
- All adult animals, including those dying during the study, were subjected to a full external and internal examination, and any macroscopic abnormalities were recorded.

HISTOPATHOLOGY / ORGAN WEIGHTS
Organ Weights: The epididymides and testes were removed from terminal kill adult males dissected free from fat and weighed before fixation. No other organs were weighed.
Samples of the following tissues were preserved from all animals from each dose group, in buffered 10% formalin, except where stated: Coagulating gland, Prostate, Epididymides**, Seminal vesicles, Ovaries, Testes**, Mammary gland (females only), Uterus/Cervix, Pituitary, Vagina.
** = preserved in Bouin’s fluid then transferred to 70% Industrial Methylated Spirits (IMS) approximately forty-eight hours later.

The tissues from control and high dose animals, any animals dying during the study, and any animals which failed to mate or did not achieve a pregnancy were prepared as paraffin blocks, sectioned at a nominal thickness of 5 μm and stained with Haematoxylin and Eosin for subsequent microscopic examination. In addition, sections of testes and epididymides from all control and 500 mg/kg bw/day males were also stained with Periodic Acid-Schiff (PAS) stain and examined.
Postmortem examinations (offspring):
SACRIFICE
- Surviving offspring were terminated via intracardiac overdose of sodium pentobarbitone. These animals were subjected to postmortem examinations (macroscopic examination) on Day 5 post partum.

GROSS NECROPSY
- Gross necropsy consisted of a full external and internal examination.
Statistics:
The following parameters were subjected to statistical analysis:
Body weight and body weight change
Food consumption for females during gestation and lactation
Pre-coital interval and gestation length
Litter size and litter weights
Sex ratio
Corpora lutea and implantation sites
Implantation losses and viability indices
Offspring body weight and body weight change
Offspring surface righting
Adult absolute and body weight-relative organ weights (Males)
See Section "Any other information on materials and methods incl. tables" for more information on statistical procedures.
Reproductive indices:
Mating Performance and Fertility:
The following parameters were calculated from the individual data during the mating period of the parental generation:
i) Pre-coital Interval
Calculated as the time elapsing between initial pairing and the observation of positive evidence of mating.
ii) Fertility Indices
For each group the following were calculated:
Mating Index (%) = (number of animals mated/number of animals paired) x 100
Pregnancy Index (%) = (number of pregnant females/number of animals mated) x 100

Gestation and Parturition Data:
The following parameters were calculated from individual data during the gestation and parturition period of the parental generation:
i) Gestation Length
Calculated as the number of days of gestation including the day for observation of mating and the start of parturition.
ii) Parturition Index
The following was calculated for each group:
Parturition Index (%) = (number of females delivering live offspring/number of pregnant females) x 100
Offspring viability indices:
The standard unit of assessment was considered to be the litter, therefore values were first calculated for each litter and the group mean was calculated using their individual litter values. Group mean values included all litters reared to termination (Day 5 of age).
i) Implantation Losses (%)
Group mean percentile pre-implantation and post-implantation loss were calculated for each female/litter as follows:
% pre–implantation loss = [(number of corpora lutea-number of implantation sites)/number of corpora lutea] x100
% post–implantation loss = [(number of implantation sites - total number of offspring born)/number of implantation sites] x 100
ii) Live Birth and Viability Indices
The following indices were calculated for each litter as follows:
Live Birth Index (%) = (number of offspring alive on Day 1/number of offspring born) x 100
Viability Index (%) = (number of offspring alive on Day 4/ number of offspring alive on Day 1) x 100
iii) Sex Ratio (% males)
Sex ratio was calculated for each litter value on Days 1 and 4 post partum, using the following formula: (number of male offspring/total number of offspring) x 100

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Two females and one male treated with 1000 mg/kg bw/day were killed in extremis on Day 3 of treatment due to clinical signs that exceeded the severity limits for this type of study. The remaining animals of either sex treated with 1000 mg/kg bw/day showed either ataxia, lethargy, noisy respiration, pilo-erection or prostration during the first three days of treatment only and subsequently the dose level was reduced. Following the reduction of the high dose level (decreased to 500 mg/kg bw/day starting on Day 5 of the study after one day non-dosed), animals of either sex did not show any toxicologically significant clinical observations. The only clinical sign at 250 mg/kg bw/day was salivation in males, which was considered to be of no toxicological importance given that this is often observed following administration of unpalatable formulations. There were no clinical observations in animals at 60 mg/kg bw/day.
Mortality:
mortality observed, non-treatment-related
Description (incidence):
Two females and one male when treated at 1000 mg/kg bw/day were killed in extremis on Day 3 prior to the reduction of this dose level to 500 mg/kg bw/day. There were no further unscheduled deaths.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Body weight reductions were evident for the 1000 mg/kg bw/day animals of either sex during the first week of the study. Following the reduction of the high dose level to 500 mg/kg bw/day body weight there was recovery and body weight development was comparable to controls. Overall bodyweight gain was reduced in the high dose group compared to control in both sexes.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Food consumption and food efficiency were reduced for the 1000 mg/kg bw/day animals when compared to the controls. Recovery was evident thereafter following the reduction of the high dose level to 500 mg/kg bw/day. There was no body weight, food consumption, or food efficiency changes at 250 or 60 mg/kg bw/day.
Food efficiency:
effects observed, treatment-related
Description (incidence and severity):
Food consumption and food efficiency were reduced for the 1000 mg/kg bw/day animals when compared to the controls. Recovery was evident thereafter following the reduction of the high dose level to 500 mg/kg bw/day. There was no body weight, food consumption, or food efficiency changes at 250 or 60 mg/kg bw/day.
Water consumption and compound intake (if drinking water study):
no effects observed
Description (incidence and severity):
Daily visual inspections of water bottles did not reveal any overt changes.
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
No toxicologically significant microscopic abnormalities were detected for terminal kill adults.
There were adverse abnormalities detected during microscopic examinations in the three high dose animals killed in extremis at the beginning of the study. Ulcerations and inflammation with submucosal edema or focal hyperkeratosis were recorded in the forestomach of those animals.
There were no treatment related abnormalities recorded in reproductive organs during microscopic examinations.
The assessment of the integrity of the spermatogenetic cycle did not provide any evidence of impaired spermatogenesis. All findings recorded were within the range of normal background alteration.
Histopathological findings: neoplastic:
no effects observed

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
no effects observed
Description (incidence and severity):
Mating: There were no treatment related effects detected on mating performance in treated animals when compared to controls. All animals mated within the first five days of pairing (i.e. at the first oestrus opportunity). One control female did not mate due to no apparent vaginal opening. In isolation and as this finding occurred in a control female only, it is considered fortuitous. There were no treatment related effects detected in corpora lutea and implantation counts or sex ratio in any treated groups when compared to control females/litters.

Fertility: In total 8, 9, 10 and 6 females from the control, 60, 250, and 500 mg/kg bw/day dose groups respectively, gave birth to a live litter and successfully reared young to Day 5 of age. There were no treatment related effects detected on fertility in treated animals when compared to controls. Two females treated at the high dose level and one control female did not achieve pregnancy following evidence of mating. No correlating histopathology was evident in the female reproductive organs, however, the male partners for these females showed testicular tubular atrophy and/or aspermia or oligospermia in the epididymides. Both microscopic findings could be correlated macroscopically to small and flaccid testes and small epididymides. In addition, in one high dose male multinucleated spermatid giant cells were recorded. All these findings are naturally occurring background changes in rats and therefore, are considered not to be related to treatment.

Gestation Lengths: There were no treatment related effects detected on gestation length. There was a slightly longer gestation length evident at 500 mg/kg bw/day in comparison to control females. In the control group the majority of females showed gestation lengths of 22 ½ days whilst at 500 mg/kg bw/day the majority of females showed gestation lengths of 23 ½ days, however, all the individual values were within the normal ranges for rats of the strain and age used in this study (21.8 to 23.5). In the view of this fact the effect is considered not related to treatment.

Effect levels (P0)

Key result
Dose descriptor:
NOAEL
Effect level:
250 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
body weight and weight gain

Target system / organ toxicity (P0)

Key result
Critical effects observed:
no

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Description (incidence and severity):
There were no offspring clinical observations that were attributable to the test item.
Description (incidence and severity):
On Days 1 and 4 post partum, the number of live offspring per litter was slightly lower at the 500 mg/kg bw/day dose level compared to controls; however, this finding was not statistically significant. This was partially reflected by a statistically significant lower live birth index at the 500 mg/kg bw/day dose level compared to controls (P<0.05). By Day 4 post partum, 3 of the 6 litters had lost more than 3 offspring after birth at the 500 mg/kg bw/day dose level. In contrast, there were no litters in the controls or other treatment groups that lost more than 1 offspring in the same time period.
Consistent with the observed slight decrease in litter size, the number of offspring at the 500 mg/kg bw/day dose level that were noted to be found dead or missing was higher than those reported at other dose levels. Incidence of clinical signs was slightly higher in litters from females treated with 500 mg/kg bw/day when compared to control litters. The number of pups that were reported to be “found dead” or “missing” was slightly higher at the 500 mg/kg bw/day dose level compared to the others, with a reported incidence of 2, 5, 2 and 12 for the control, 60, 250 and 500 mg/kg bw/day dose groups, respectively. These findings are consistent with the slightly smaller post partum litter sizes reported at the 500 mg/kg bw/day dose level. There were no other clinical signs that were indicative of a treatment related effect.
No treatment related effects were evident in sex ratio.
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
There were no treatment related effects in total litter weights, offspring body weight and body weight changes.
Mean litter weight values were slightly reduced in 500 mg/kg bw/day litters due to smaller litter size values recorded in this treatment group. In view of the fact that mean offspring weight values at Day 1 and 4 assessments were comparable to controls this finding is considered not related to treatment.
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Description (incidence and severity):
No obvious adverse macroscopic abnormalities were detected for interim death or terminal kill offspring. The incidental findings observed were those occasionally observed in reproductive studies of this type and were considered to be unrelated to toxicity of the test item.
Histopathological findings:
not examined

Developmental neurotoxicity (F1)

Behaviour (functional findings):
effects observed, non-treatment-related
Description (incidence and severity):
Surface righting mean values appeared slightly reduced in 500 mg/kg bw/day litters when compared to control values. This effect did not reach statistical significance and a true dose related response was not evident. All values were also within the normal ranges for the strain used on this study, therefore, the intergroup difference was considered not to be treatment related.

Developmental immunotoxicity (F1)

Developmental immunotoxicity:
not examined

Effect levels (F1)

open allclose all
Key result
Dose descriptor:
LOAEL
Generation:
F1
Effect level:
500 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
viability
Key result
Dose descriptor:
NOEL
Generation:
F1
Effect level:
250 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No effect observed

Target system / organ toxicity (F1)

Key result
Critical effects observed:
no

Overall reproductive toxicity

Key result
Reproductive effects observed:
no

Any other information on results incl. tables

None

Applicant's summary and conclusion

Conclusions:
Under the test conditions, the NOAEL for adult toxicity is 250 mg/kg bw/day and NOAEL for reproductive/developmental toxicity is 250 mg/kg bw/day in rats.
Executive summary:

In a Reproduction / Developmental Toxicity Screening Test conducted according to OECD Guideline 421 and in compliance with GLP, test item was administered to groups of Wister HanTM: RccHanTM: WIST strain rats (10/sex/dose) at 0, 60, 250, 1000/500 mg/kg bw/day by oral (gavage) for up to eight weeks (including a two week maturation phase, pairing, gestation and early lactation for females), at dose levels of 60, 250 and 500 mg/kg bw/day (Male: Days 1-43 (mating on Days 15/16); Female: Days 1 to post partum Day 5 (mating on Days 15/16)). A control group of ten males and ten females was dosed with vehicle alone (Arachis oil BP). Pairing of animals within each dose group was undertaken on a 1 male: 1 female basis within each treatment group following at least fourteen days of treatment, with females subsequently being allowed to litter and rear their offspring to Day 5 of lactation. Clinical signs, body weight change, dietary intake and water consumption were monitored during the study. All animals were subjected to a gross necropsy examination, selected organs were weighed and histopathological evaluation of selected tissues was performed. The clinical condition of offspring, litter size and survival, sex ratio and offspring bodyweight were assessed and macroscopic pathology investigations were undertaken.

 

Two females and one male when treated at 1000 mg/kg bw/day were killed in extremis on Day 3 prior to the reduction of this dose level to 500 mg/kg bw/day. The remaining animals of either sex treated with 1000 mg/kg bw/day showed either ataxia, lethargy, noisy respiration, pilo-erection or prostration during the first three days of treatment only. There were no further unscheduled deaths. Following the reduction of the high dose level (decreased to 500 mg/kg bw/day starting on Day 5 of the study after one day non-dosed), animals of either sex did not show any toxicologically significant clinical observations. Body weight reductions were evident for the 1000 mg/kg bw/day animals of either sex during the first week of the study. Following the reduction of the high dose level to 500 mg/kg bw/day body weight development was comparable to controls. Overall bodyweight gain was reduced in both males and females in the high dose group when compared to the control group. Food consumption and food efficiency were reduced for the 1000 mg/kg bw/day animals when compared to the controls. Recovery was evident thereafter following the reduction of the high dose level to 500 mg/kg bw/day. There were no treatment related effects detected on mating performance, fertility and gestation length in treated animals when compared to controls.

 

Litter Responses:

Offspring Litter Size, Sex Ratio and Viability: No treatment related effects were evident in sex ratio. On Days 1 and 4 post partum, the number of live offspring per litter was slightly lower at the 500 mg/kg bw/day dose level compared to controls; however, this finding was not statistically significant. This was partially reflected by a statistically significant lower live birth index at the 500 mg/kg bw/day dose level compared to controls (P<0.05). By Day 4 post partum, 3 of the 6 litters had lost more than 3 offspring after birth at the 500 mg/kg bw/day dose level. In contrast, there were no litters in the controls or other treatment groups that lost more than 1 offspring in the same time period.

Offspring Growth and Development: No treatment related effects were detected in offspring growth and development.

Offspring Observations: Consistent with the observed slight decrease in litter size, the number of offspring at the 500 mg/kg bw/day dose level that were noted to be found dead or missing was higer than those reported at other dose levels. There were no other offspring clinical observations that were attributable to the test item.

 

Pathology: The 1000 mg/kg bw/day male and two 1000 mg/kg bw/day females that were killed in extremis had a raised limiting ridge in the stomach at necropsy, considered as evidence of local irritation. No toxicologically significant macroscopic, microscopic abnormalities and changes in organ weights were detected for terminal kill adults and no obvious adverse macroscopic findings were detected for offspring.

Under the test conditions, the NOAEL for adult toxicity is 250 mg/kg bw/day and NOAEL for reproductive/developmental toxicity is 250 mg/kg bw/day in rats.