Registration Dossier

Diss Factsheets

Toxicological information

Developmental toxicity / teratogenicity

Currently viewing:

Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
25 March - 20 May 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: This study has been performed according to OECD and/or EC guidelines and according to GLP principles.
Cross-reference
Reason / purpose for cross-reference:
reference to same study

Data source

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

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
other: Organisation of Economic Co-operation and Development (OECD) Guidelines for Testing of Chemicals, Guideline 422, Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test, March 1996
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: The United States Environmental Protection Agency (EPA) Health Effects Test Guidelines, OPPTS 870.3650, Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test, July 2000.
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: OECD Guidelines for Testing of Chemicals, Guideline 421, Reproduction/Developmental Toxicity Screening Test, July 1995.
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: The United States EPA Health Effects Test Guidelines, OPPTS 870.3550, Reproduction/Developmental Toxicity Screening Test, July 2000.
Deviations:
no
Principles of method if other than guideline:
In addition, the procedures described in the report essentially conform to the following guidelines:
- Commission regulation (EC) No 440/2008 Part B: Methods for the Determination of Toxicity and other Health Effects; B.7: "Repeated Dose (28 days) Toxicity (oral)". Official Journal of the European Union No. L142, May 2008.
- OECD Guidelines for Testing of Chemicals, Guideline 407, Repeated Dose 28-day Oral Toxicity Study in Rodents, October 2008.
- The United States EPA Health Effects Test Guidelines, OPPTS 870.3050, Repeated dose 28-day oral toxicity study in rodents, July 2000.
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Amines, C36-alkylenedi-
EC Number:
273-282-8
EC Name:
Amines, C36-alkylenedi-
Cas Number:
68955-56-6
Molecular formula:
Not applicable UVCB substance
IUPAC Name:
(1E,19E)-10,11-dioctylicosa-1,19-diene-1,20-diamine
Test material form:
other: Slightly viscous amber liquid
Details on test material:
- Name of test material (as cited in study report): Amines, C36-alkylenedi-
- Substance type: Slightly viscous amber liquid
- Physical state: liquid
- Purity: 98.2%
- Lot/batch No.: 0000436391
- Expiration date of the lot/batch: 24 January 2021
- Storage condition of test material: At room temperature in the dark under nitrogen
- pH: 9.9-9.9
- Density: 0.885 g/cm3 at 20ºC
- Volatile: No
- Test substance handling: Flush container with nitrogen after handling
- Solubility in Water: No

Test animals

Species:
rat
Strain:
other: Crl:WI(Han)
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany.
Nulliparous and nonpregnant females and untreated animals were used at initiation of the study.
- Age at study initiation: Approximately 11 weeks.
- Weight at study initiation: mean weight at start of treatment was 293 gr (males) or 201 gr (females).
- Fasting period before study: no
- Housing:
Pre-mating: Animals were housed in groups of 5 animals/sex/cage in Macrolon cages.
Mating: Females were caged together with males on a one-to-one-basis in Macrolon cages.
Post-mating: Males were housed in their home cage with a maximum of 5 animals/cage. Females were individually housed in Macrolon cages.
General: Sterilised sawdust as bedding material and paper as cage enrichment were supplied.
- Diet: Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).
- Water: Free access to tap water.
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
Environmental controls for the animal room were set to maintain 18 to 24°C, a relative humidity of 40 to 70%, approximately 15 room air changes/hour, and a 12-hour light/12-hour dark cycle.
Temporary deviations from the minimum level of daily mean relative humidity occurred. Laboratory historical data do not indicate an effect of the deviations.

IN-LIFE DATES: From: 25 March - 20 May 2012

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
propylene glycol
Details on exposure:
- Method of formulation: Formulations (w/w) were prepared daily within 6 hours prior to dosing and were homogenized to a visually acceptable level. Adjustment was made for specific gravity of the vehicle and density of the test substance and vehicle. No correction was made for the purity of the test substance.
- Storage conditions of formulations: At ambient temperature.
- Justification for use and choice of vehicle: Based on trial formulations performed at WIL Research Europe B.V. and on information provided by the Sponsor.
- Dose volume: 5 mL/kg body weight. Actual dose volumes were calculated according to the latest body weight.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analyses were conducted on a single occasion after the treatment phase, since a validated method (Project 498062) was not yet available during the in-life phase. Samples of formulations prepared in an identical manner as during the in-life phase were analyzed for homogeneity (highest and lowest concentration) and accuracy of preparation (all concentrations). Stability in vehicle over 6 hours at room temperature was also determined (highest and lowest concentration). The accuracy of preparation was considered acceptable if the mean measured concentrations were 85-115% of the target concentration for Group 4 or 85-115% of the target concentration for Groups 2 and 3. Homogeneity was demonstrated if the coefficient of variation was ≤ 10%. Formulations were considered stable if the relative difference before and after storage was maximally 10%.

Accuracy of preparation results:
The concentrations analysed in the formulations of Group 3 and Group 4 were in agreement with target concentrations (i.e. mean accuracies between 85% and 115%).
For the formulation of Group 2, the mean accuracy was above the target concentration (i.e. 117% of target). Since the deviation above 115% was relatively small and since it was known from the method validation (see NOTOX project 498062) that recoveries can vary more than normal due to the properties of the substance, this was accepted.
A small response at the retention time of the test substance was observed in the chromatograms of the Group 1 formulation prepared after in-life phase. It was not considered to derive from the formulation since a similar response was obtained in the analytical blanks.

Homogeneity results:
The formulations of Group 2 and Group 4 were homogeneous (i.e. coefficient of variation ≤ 10%).

Stability results:
Formulations at the entire range were stable when stored at room temperature under normal laboratory light conditions for at least 6 hours.
Details on mating procedure:
- M/F ratio per cage:1/1 (one female was cohabitated with one male of the same treatment group, avoiding sibling mating (Charles River supplied non-litter mates).
- Age at mating of the mated animals in the study: Approximately 13 weeks
- Length of cohabitation: A maximum of 14 days was allowed for mating.
- Proof of pregnancy: Detection of mating was confirmed by evidence of sperm in the vaginal lavage, by the appearance of an intravaginal copulatory plug and/or by determination of the estrous stage. This day was designated Day 0 post-coitum. Once mating had occurred, the males and females were separated.
- \\\evt toevoegen: After 14 days of unsuccessful pairing \\ one female who had not shown evidence of mating was separated from her male.
- Further matings after two unsuccessful attempts: no \\zie report
- After successful mating each pregnant female was caged individually in Macrolon cages (MIII type, height 18 cm).
- Any other deviations from standard protocol: no
Duration of treatment / exposure:
Males were exposed for 29 days, i.e. 2 weeks prior to mating, during mating, and up to termination. Females were exposed for 43-57 days, i.e. during 2 weeks prior to mating, during mating, during post-coitum, and during at least 4 days of lactation. Four females (all of different Groups) were not dosed during littering.

Pups were not treated directly, but were potentially exposed to the test substance in utero and through lactational transfer.
Frequency of treatment:
Once daily, 7 d/w
On Day 36, one female of Group 3 was dosed twice (within 30 minutes; 88% of the intended dose at the first dosing, and 12% of the intended dose at the second dosing). A clinical observation was conducted only after the first dosing. The animal eventually received the correct volume and dose. Dosing an animal twice within 30 minutes is considered to have no adverse effect on the outcome of the study given its incidental occurrence, and since oral gavage is not an invasive procedure. The observation was conducted after the animal received the larger part of the dose (i.e. after the first dose), and since no peak effect of occurrence of clinical signs was noted in the dose range finding study (NOTOX Project 498061), this was considered sufficient for the purpose of this study.
One female of Group 1 was dosed on the day of necropsy (lactation Day 7). This did not adversely affect any subsequent examinations conducted for this animal. This animal was not selected for clinical pathology determinations and was hence not fasted overnight prior to necropsy/dosing.
Duration of test:
Males: 29 days
Females: 43-57 days
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 50, 150, 500 mg/kg bw/day
Basis:
actual ingested
No. of animals per sex per dose:
10; an extra 5 males in Groups 1 and 4 were allowed 2 weeks of recovery.

Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: In consultation with the sponsor and based on the results of the dose range finding study (NOTOX Project 498060: Based on the results of this range finding study, dose levels for the main study were 50, 150 and 500 mg/kg body weight. No clinical signs were observed at the highest selected dose for the main study (500 mg/kg). Therefore, clinical observations in the main study were conducted immediately after dosing, and functional observation tests were conducted after dosing at no specific time point, but within a similar time period after dosing for the respective animals.)
- Parturition: The females were allowed to litter normally. Day 1 of lactation was defined as the day when a litter was found completed (i.e. membranes, placentas cleaned up, nest built up and/or feeding of pups started). Females that were littering were left undisturbed.
- Identification of pups: On Day 1 of lactation, all pups were individually identified by means of subcutaneous injection of Indian ink.
- Selection of animals for selected measurements: 5 animals/sex/group were randomly selected at allocation for functional observations, clinical pathology, macroscopic examination (full list), organ weights (full list) and histopathology (see also respective paragraphs). Only females with live offspring were selected.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS:
- Time schedule: At least twice daily. The circumstance of any death was recorded in detail.

DETAILED CLINICAL OBSERVATIONS:
- Time schedule: Daily detailed clinical observations were made in all animals, within 0-30 minutes after dosing between Days 1-18, and within 0-15 minutes after dosing from day 19 onwards. Once prior to start of treatment and at weekly intervals during the treatment period this was also performed outside the home cage in a standard arena. The time of onset, grade and duration of any observed sign was recorded. Signs were graded for severity.
On the following days, clinical observations were conducted later than protocolled: Day 15, females (start of observations approximately 6 minutes later); Day 17, males (start of observations approximately 3 minutes later); Daily from Day 18 onwards: maximum 23 minutes later. The slightly later time point of observations did not adversely affect the interpretation of the study results, given the absence of a peak effect of occurrence of clinical signs (based on the dose range finding study (NOTOX Project 498061).
No arena observations were performed in weeks 8 and 9 of the study. The daily observations for clinical signs consist of a series of extensive examinations including the type of observations that are also assessed in the arena observations, and as such, the likelihood of having missed any relevant observations is minimal.

BODY WEIGHT:
- Males and females were weighed on the first day of exposure and weekly thereafter. Mated Main females were weighed on Days 0, 4, 7, 11, 14, 17 and 20 post-coitum and during lactation on Days 1 and 4.

FOOD CONSUMPTION:
- Weekly, except for Main males and Main females which were housed together for mating and for Main females without evidence of mating. Food consumption of mated Main females was measured on Days 0, 4, 7, 11, 14, 17 and 20 post-coitum and on Days 1 and 4 of lactation.

FOOD EFFICIENCY:
- (average food consumption [per animal per day]/average body weight per cage)x1000

WATER CONSUMPTION : No.
Subjective appraisal was maintained during the study, but no quantitative investigation introduced as no effect was suspected.

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY:
- Time schedule for collection of blood: immediately prior to scheduled post mortem examination.
- Anaesthetic used for blood collection: Yes (iso-flurane)
- Animals fasted: Yes (with a maximum of 20 hours). Water was provided.
- How many animals: 5 animals/sex/group
- Parameters checked were: According to test guidelines

CLINICAL CHEMISTRY:
- Time schedule for collection of blood: immediately prior to scheduled post mortem examination.
- Animals fasted: Yes (with a maximum of 20 hours). Water was provided.
- How many animals: 5 animals/sex/group
- Parameters checked were: According to test guidelines

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION:
- Time schedule for examinations: The selected Main males were tested during Week 4 of treatment and the selected females were tested during lactation (all before blood sampling). Since (potential) treatment-related findings were noted in motor activity of males at 500 mg/kg, motor activity measurements were also conducted for all Recovery males at the end of the recovery phase.
- Dose groups that were examined: all
- Battery of functions tested: According to test guidelines
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: No
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: No
- Number of late resorptions: No
Fetal examinations:
PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross abnomalies, weight gain, physical or behavioural abnormalities.

- Mortality: The numbers of live and dead pups on Day 1 of lactation and daily thereafter were determined. If possible, defects or cause of death were evaluated.
- Clinical signs: At least once daily, detailed clinical observations were made in all animals.
During the lactation period, clinical observations were not recorded for several pups and litters on one or more days. Evaluation: sufficient clinical observations were conducted to allow adequate interpretation of the study results.
- Body weights: Live pups were weighed on Days 1 and 4 of lactation.
- Sex: Sex was determined for all pups on Days 1 and 4 of lactation (by assessment of the ano-genital distance).

GROSS EXAMINATION OF DEAD PUPS
Yes, if possible, defects or cause of death were evaluated.
Statistics:
The following statistical methods were used to analyse the data:
- If the variables could be assumed to follow a normal distribution, the Dunnett-test (Dunnett, 1955) (many-to-one t-test) based on a pooled variance estimate was applied for the comparison of the treated groups and the control groups for each sex. (Ref. 1)
- The Steel-test (Miller, 1981) (many-to-one rank test) was applied if the data could not be assumed to follow a normal distribution. (Ref. 3)
- The Fisher Exact-test (Fisher, 1950) was applied to frequency data. (Ref. 2)
- The following additional methods of statistical analysis were used: Motor activity data was subjected to the Kruskal-Wallis nonparametric ANOVA test (Ref. 4) to determine intergroup differences followed by the Wilcoxon test (Ref. 5) to compare the treated groups to the control group.

All tests were two-sided and in all cases p < 0.05 was accepted as the lowest level of significance. Group means were calculated for continuous data and medians were calculated for discrete data (scores) in the summary tables. No statistical analysis was performed on histopathology findings.

References:
Ref. 1 Dunnett C.W., A Multiple Comparison Procedure for Comparing Several Treatments with a Control, J. Amer. Stat. Assoc. 50, 1096-1121 (1955).
Ref. 2 Fisher R.A., Statistical Methods for Research Workers, Oliver and Boyd, Edinburgh (1950).
Ref. 3 Miller R.G., Simultaneous Statistical Inference, Springer Verlag, New York (1981).
Ref. 4 Kruskal W.H. and Wallis W.A.. Use of ranks in one-criterion variance analysis. Journal of the American Statistical Association 47 (260): 583-621, December (1952).
Ref. 5 Wilcoxon, F. Individual comparisons by ranking methods. Biometrics, 1, 80-83 (1945).
Indices:
Reproductive indices; For each group, the following calculations were performed:
- Mating index: Number of females mated/Number of females paired x 100
- Fertility index: Number of pregnant females/Number of females paired x 100
- Conception index: Number of pregnant females/Number of females mated x 100
- Gestation index: Number of females bearing live pups/Number of pregnant females x 100
- Duration of gestation: Number of days between confirmation of mating and the beginning of parturition

Offspring indices:
- Percentage live males at First Litter Check: Number of live male pups at First Litter Check/Number of live pups at First Litter Check x 100
- Percentage live females at First Litter Check: Number of live female pups at First Litter Check/Number of live pups at First Litter Check x 100
- Percentage of postnatal loss Days 0-4 of lactation: Number of dead pups on Day 4 of lactation/Number of live pups at First Litter Check x 100
- Viability index: Number of live pups on Day 4 of lactation / Number of pups born alive x 100

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Details on maternal toxic effects:
MORTALITY
No mortality occurred during the study period that was considered to be related to treatment with the test substance. One female at 500 mg/kg was found dead on Day 14 of the post-coitum period. Macro- and microscopic findings observed for this animal indicated that its death was gavage-related. In addition, in the absence of similar findings or signs of moribundity among other animals of this dose group, this death was considered to be unrelated to toxicity of the test substance.

CLINICAL SIGNS
No clinical signs of toxicity were noted during the observation period. Salivation seen after dosing among all animals at 500 mg/kg and at lower incidence at 150 mg/kg during the treatment period was considered to be a physiological response rather than a sign of systemic toxicity considering the nature and minor severity of the effect and its time of occurrence (i.e. after dosing). Incidental findings that were noted included alopecia, a broken tail apex and rales. One female at 50 mg/kg showed piloerection, hunched posture and a lean appearance during two weeks prior to scheduled necropsy. Piloerection was also noted for one female each at 50 and 500 mg/kg. At the incidence observed, these were considered signs of no toxicological relevance.

FUNCTIONAL OBSERVATIONS
No toxicologically significant effects on motor activity were noted. Hearing ability, pupillary reflex, static righting reflex and grip strength were normal in all animals. Motor activity of males at 500 mg/kg at the end of the treatment period was slightly higher than control animals, achieving a level of statistical significance for ambulation counts. Motor activity of females at 500 mg/kg during the lactation period showed the opposite effect, i.e. being lower than control animals, achieving a level of statistical significance for total movements and ambulation counts. At the end of the recovery period (assessed for males only), motor activity of males at 500 mg/kg was similar to control levels. Since the change in motor activity was reversible in males, the motor activity habituation profile appeared similar between the groups, clinical observations revealed no supportive findings in both sexes and functional observation tests were normal, these changes in motor activity were considered not to be of toxicological relevance.

BODY WEIGHTS
Body weights and body weight gain of treated animals remained in the same range as controls over the treatment period.

FOOD CONSUMPTION
Food consumption before or after correction for body weight was similar between treated and control animals during the treatment period. One female at 50 mg/kg showed a notable reduction in food intake over Days 11-20 of the post-coitum period, coinciding with occurrence of clinical signs (see respective paragraph). Given the incidental nature of this change, this was considered to be of no toxicological relevance.

HAEMATOLOGY
At the end of the treatment period, a higher mean white blood cell count occurred in males and females at 500 mg/kg (not statistically significant for females), which was essentially due to a higher mean neutrophil count (relative and absolute counts increased with statistical significance). At the end of the recovery period (assessed for males only), the mean white blood cell count and mean relative and absolute neutrophil count remained increased over the control mean.

CLINICAL BIOCHEMISTRY
No toxicologically relevant changes occurred in clinical biochemistry parameters of treated rats.

MACROSCOPIC EXAMINATION
Necropsy did not reveal any toxicologically relevant alterations. Macroscopic findings in the female at 500 mg/kg found dead on Day 14 of the post-coitum period consisted of gray-white discolouration of the left side of the lungs and reddish fluid in the thoracic cavity. The nature of these findings, and absence of similar findings among other animals of the same group indicate that these were gavage-related. The incidence of other necropsy findings among surviving control and treated animals at the end of the treatment and recovery period was within the background range of findings that are encountered among rats of this age and strain, and did not show a dose-related incidence trend. These necropsy findings were therefore considered to be of no toxicological relevance.

ORGAN WEIGHTS
No toxicologically relevant changes were noted in absolute and relative organ weights.

MICROSCOPIC EXAMINATION
The following microscopic findings were considered to be related to treatment with the test substance:
- Duodenum: foamy macrophage foci in the lamina propria were present in 1/5 500 mg/kg (minimal) treated Main Group female rats.
- Jejunum: foamy macrophage foci in the lamina propria were present in 1/5 500 mg/kg (minimal) treated female rats.
- Ileum: foamy macrophage foci in the lamina propria were present in 1/5 150 mg/kg (1 minimal) and in 4/5 500 mg/kg (4 minimal) treated female rats.
- Mesenteric lymph node:
- Macrophage foci were present at increased incidence and severity in 5/5 50 mg/kg (4 slight, 1 moderate), in 5/5 150 mg/kg (1 slight, 3 moderate, 1 marked) and in 5/5 500 mg/kg (1 minimal, 1 slight, 3 moderate) treated female rats (no foci in control females).
- Fibrosis was present in 2/5 500 mg/kg treated female rats (1 minimal, 1 slight)
- Sinus histiocytosis was present in 3/5 50 mg/kg treated female (3 minimal) rats, in 5/5 female rats at 150 mg/kg (2 minimal, 3 slight), and in 5/5 female rats at 500 mg/kg (2 slight, 3 moderate).
There were no morphological findings in the reproductive organs of females which could be attributed to the test item.

Effect levels (maternal animals)

open allclose all
Dose descriptor:
NOAEL
Effect level:
50 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEL
Effect level:
>= 500 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects. Remark: Only early postnatal pup development parameters were examined including body weight, post-natal loss, sex ratio, clinical signs, body weight and external macroscopy.

Details on embryotoxic / teratogenic effects:
Early postnatal pup development
Number of dead and living pups at first litter check, postnatal loss, viability index and sex ratio were unaffected by treatment, and clinical signs, body weight and external macroscopy did not reveal treatment-related findings.

Mortality
One pup of the control group, and two pups each at 50, 150 and 500 mg/kg/day were found dead or missing during lactation. Pups missing were most likely cannibalised. No toxicological relevance was attributed to these dead/missing pups since the mortality incidence did not show a dose-related trend and remained within the range considered normal for pups of this age.

Clinical signs
Incidental clinical symptoms of pups consisted of scabbing of the head, back or right hind leg, lean appearance and absence of milk in the stomach. The nature and incidence of these clinical signs remained within the range considered normal for pups of this age, and were therefore considered to be of no toxicological relevance.

Body weights
Body weights of pups were considered to have been unaffected by treatment.

Macroscopy
Incidental macroscopic findings of pups that were found dead included absence of milk in the stomach, lean appearance and autolysis. The nature and incidence of these findings remained within the range considered normal for pups of this age, and were therefore considered to be of no toxicological relevance.

There was no evidence of teratogenic effects based on the absence of relevant clinical signs and external macroscopic findings.

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Any other information on results incl. tables

REPRODUCTIVE DATA
No toxicologically relevant effects on number of corpora lutea and implantation sites were noted.

GESTATION
No toxicologically relevant effects on gestation index and duration, parturition, maternal care and early postnatal pup development (mortality, clinical signs, body weight and macroscopy) were observed. One control female did not deliver but was found to be pregnant based on presence of implantation sites. The number of non-pregnant females (one at each dose group) showed no dose-related incidence. One pregnant female at 500 mg/kg died before delivery, which was considered to be gavage-related. The statistically significant higher duration of gestation at 50 mg/kg occurred in the absence of a dose-related trend and was slight in nature. No toxicological significance was ascribed to this change.

PARTURITION/MATERNAL CARE
No signs of difficult or prolonged parturition were noted among the pregnant females. Examination of cage debris of pregnant females revealed no signs of abortion or premature birth. No deficiencies in maternal care were observed.

Applicant's summary and conclusion

Conclusions:
In conclusion, treatment with Amines, C36-alkylenedi-, by oral gavage in male and female Wistar Han rats at dose levels of 50, 150 and 500 mg/kg revealed parental toxicity at 50 mg/kg and higher. At 50 mg/kg, findings were confined to macrophage foci and sinus histiocytosis in the mesenteric lymph nodes (minimal to moderate degree). At 150 and/or 500 mg/kg, these findings were noted at higher incidence and/or severity, along with foamy macrophages in the lamina propria of the duodenum and/or ileum, and fibrosis (two females at 500 mg/kg) in the mesenteric lymph nodes, and higher white blood cell (neutrophil) counts at 500 mg/kg. The nature of changes at 150 and 500 mg/kg (in particular necrosis and fibrosis in the mesenteric lymph nodes, albeit occurring at low incidence), indicate that these effects are to be considered adverse in toxicological terms at those dose levels.

No developmental toxicity was observed for treatment up to 500 mg/kg body weight/day.

Based on these results, the following No Observed Adverse Effect Levels (NOAEL) were derived: Parental NOAEL: 50 mg/kg, Developmental NOAEL: at least 500 mg/kg.