Registration Dossier

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

500 mg/kg bw/day

Study duration:

subacute

Species:

rat

Endpoint conclusion:

no study available

Endpoint conclusion:

no study available

GUIDELINE

Repeated dose oral toxicity was considered in a study designed to investigate the systemic toxicity and potential adverse effects of the test item on reproduction (including offspring development); it also assesses the ability of the animals to recover from any toxicity following the withdrawal of treatment. The study was compatible with the requirements of the OECD Guidelines for Testing of Chemicals No. 422 “Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test” (adopted 22 March 1996).

METHODS

The test item was administered by gavage to three groups, each of twelve male and twelve female Wistar Han:RccHan:WIST strain rats, for approximately six weeks (including a two week pre-pairing phase, pairing, gestation and early lactation for females), at dose levels of 100, 250 and 500 mg/kg bw/day. A control group of twelve males and twelve females was dosed with vehicle alone (Arachis oil BP) over the same treatment period. Two recovery groups, each of five males and five females, were treated with the high dose (500 mg/kg bw/day) or the vehicle alone for forty-two consecutive days and then maintained without treatment for a further fourteen days.

Clinical signs, behavioral assessments, body weight change and food and water consumption were monitored during the study.

Pairing of non-recovery animals within each dose group was undertaken on a one male: one female basis within each treatment group on Day 15 of the study, with females subsequently being allowed to litter and rear their offspring to Day 5 of lactation.

During the lactation phase, daily clinical observations were performed on all surviving offspring, together with litter size and offspring weights and assessment of surface righting reflex. Extensive functional observations were performed on five selected non-recovery males from each dose group after the completion of the pairing phase, and for five selected parental females from each dose group on Day 4 post partum. Urinalysis was performed on five non-recovery males per dose group during the final week of treatment and five non-recovery males and females from each dose group were selected for hematology and blood chemistry assessments prior to termination. Surviving adult non-recovery males were terminated on Day 43 or Day 44, with all females and offspring being killed on Day 5 post partum. Any female which did not produce a pregnancy was terminated on or after Day 25 post coitum. All animals were subjected to a gross necropsy examination and histopathological evaluation of selected tissues was performed. Following forty-two days of treatment, recovery group animals were maintained without treatment for a further fourteen days. Urinalysis was performed on all recovery group males during the final week of the treatment period. In addition, hematological and blood chemical assessments were performed on all recovery group animals at the end of the treatment-free period. These animals were then subjected to a gross necropsy and histopathological examinations of selected tissues was performed.

RESULTS

Mortality: At 100 mg/kg bw/day, one male died during the blood sampling procedure on Day 42; this death was considered to be incidental and unrelated to treatment.

Clinical observations: Treatment at 500 mg/kg bw/day was associated with increased post-dosing salivation in all seventeen male and seventeen female; ten males and eight females also showed noisy respiration during the study. Increased post-dosing salivation was also observed for five males at 100 mg/kg bw/day and the twelve males and eight females at 250 mg/kg bw/day.

Behavioural assessment: There were no treatment-related changes in the behavioural parameters at 100, 250 or 500 mg/kg bw/day.

Functional performance tests: There were no changes in functional performance considered to be related to treatment at 100, 250 or 500 mg/kg bw/day.

Sensory reactivity assessments: There were no inter-group differences in sensory reactivity scores.

Body weight: At 500 mg/kg bw/day, males showed lower body weight gain during the first week of treatment, thereafter there was recovery of body weight gain although body weight gain still tended to be lower than control during the treatment period. Body weight gain of males was slightly superior to control during the recovery period. For females at 500 mg/kg bw/day, lower body weight gain was also observed during the first week of treatment. There was no subsequent effect of treatment on body weight gain during the remainder of the study, including gestation, lactation and recovery phases. At 250 mg/kg bw/day, males showed lower body weight gain during the first week of treatment; body weight gain was considered to be unaffected by treatment for the remaining treatment period and the recovery period. At both sexes at 100 mg/kg bw/day and females at 250 mg/kg bw/day, body weight gain was considered to be unaffected by treatment.

Food consumption: At 500 mg/kg bw/day, food consumption for both sexes was slightly lower than control during the first week of treatment; thereafter food intake was unaffected by treatment for the remainder of the study including gestation, lactation and recovery phases. At 100 and 250 mg/kg bw/day, food consumption for both sexes was considered to be unaffected by treatment throughout the study.

Food consumption efficiency: At 500 mg/kg bw/day, food conversion efficiency for both sexes was slightly lower thancontrol during the first week of treatment. At 100 and 250 mg/kg bw/day, food conversion efficiency for both sexes was considered to be unaffected by treatment.

Water consumption: Water consumption for both sexes was unaffected by treatment at 100, 250 or 500 mg/kg bw/day.

Reproductive performance (mating): Mating performance was unaffected by treatment at 100, 250 or 500 mg/kg bw/day.

Reproductive performance (fertility): Fertility was unaffected by treatment at 100, 250 or 500 mg/kg bw/day.

Reproductive performance (gestation lengths): Gestation length was unaffected by treatment at 100, 250 or 500 mg/kg bw/day.

Litter responses (offspring litter size, sex ratio and viability): The number of corpora lutea and implantations, pre- and post implantation loss, litter size, sex ratio and offspring survival was unaffected by treatment at 100, 250 or 500 mg/kg bw/day.

Litter responses (offspring growth and development): At 500 mg/kg bw/day, offspring body weight for both sexes was lower than control at Day 1 of age; surface righting performance on Day 1, litter weights on Day 1 and 4 and body weight gain to Day 4 was considered to be unaffected by maternal treatment. At 100 and 250 mg/kg bw/day, offspring body weights, body weight gain, litter weights and surface righting performance on Day 1 was unaffected by maternal treatment. Clinical signs and necropsy findings observed for offspring did not indicate any effect of maternal treatment on offspring development at 100, 250 or 500 mg/kg bw/day.

Laboratory investigations: There was considered to be no adverse effect of treatment on hematology parameters for either sex at 100, 250 or 500 mg/kg bw/day.

Blood chemistry: At 500 mg/kg bw/day, males showed higher alanine aminotransferase and total cholesterol levels and females higher aspartame aminotransferase and alanine aminotransferase levels at the end of the treatment.

Urinalysis: Analysis of urinalysis parameters for males at 100, 250 or 500 mg/kg bw/day during treatment and for males at 500 mg/kg bw/day following recovery did not indicate any effect of treatment.

Necropsy: At 500 mg/kg bw/day, raised areas in the non-glandular region of the stomach were apparent for four males and five females at the end of the treatment period, with three of the females also showing sloughing of the non-glandular/glandular region of the stomach.

Organ weights: At 250 and 500 mg/kg bw/day, absolute and body weight-relative liver weights at the end of the treatment period were statistically significantly higher than control for both sexes at the end of treatment. For males at 500 mg/kg bw/day, statistically significantly lower absolute and body weight relative seminal vesicles weights were observed at the end of the treatment period, but only two individual absolute values and one body weight relative value were below the historical control range.

Histopathology (liver): Bile duct hyperplasia was observed for the majority of animals at 250 mg/kg bw/day and all animals at 500 mg/kg bw/day. Periductal inflammation/fibrosis was present for two females at 250 mg/kg bw/day and one male and all females at 500 mg/kg bw/day. Periductal infiltration, mainly lymphocytic, was present in the majority of males and two females at 250 mg/kg/day. Centrilobular hypertrophy was present for females at 250 mg/kg bw/day and males at 500 mg/kg bw/day. Females at 500 mg/kg bw/day showed focal necrosis at a minimal level. After the recovery period the bile duct changes persisted for both sexes with a slight reduction in severity; other changes had resolved.

Histopathology (stomach): Hyperplasia of the non-glandular epithelium (mild, moderate or marked) was observed at 500 mg/kg bw/day at the end of treatment. This was still apparent at notably reduced severity after the treatment free period indicating partial recovery.

Histopathology (spleen): There was a minor increase in the severity of hemopoiesis in the spleen of females at 250 and 500 mg/kg bw/day at the end of treatment; this had resolved after the recovery period.

Histopathology (thyroid gland): Hypertrophy of the follicular epithelium was increased for both sexes at 500 mg/kg bw/day at the end of treatment; this had resolved after the recovery period.

Histopathology (Urinary bladder): Mild, diffuse urothelial hyperplasia was present for two females at 250 mg/kg bw/day and one male and three females at 500 mg/kg bw/day at the end of treatment. Evidence of recovery, with a reduction in incidence and severity without complete resolution, was apparent after the recovery period.

CONCLUSION

Treatment at 250 and 500 mg/kg bw/day was associated with adverse liver histopathology, although the toxicological significance of these findings for man is considered to be equivocal. The No Observed Adverse Effect Level (NOAEL) for systemic toxicity of the rat was considered to be 100 mg/kg bw/day.The No Observed Adverse Effect Level (NOAEL) for reproduction and the survival, growth and development of the offspring was considered to be 500 mg/kg bw/day.

Short description of key information:
The No Observed Adverse Effect Level (NOAEL) for reproduction and the survival, growth and development of the offspring was considered to be 500 mg/kg bw/day (OECD 422).

Justification for selection of Effect on fertility via oral route:
GLP guideline study.

The No Observed Adverse Effect Level (NOAEL) for reproduction and the survival, growth and development of the offspring was considered to be 500 mg/kg bw/day (OECD 422).

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

500 mg/kg bw/day

Study duration:

subacute

Species:

rat

Endpoint conclusion:

no study available

Endpoint conclusion:

no study available

GUIDELINE

Repeated dose oral toxicity was considered in a study designed to investigate the systemic toxicity and potential adverse effects of the test item on reproduction (including offspring development); it also assesses the ability of the animals to recover from any toxicity following the withdrawal of treatment. The study was compatible with the requirements of the OECD Guidelines for Testing of Chemicals No. 422 “Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test” (adopted 22 March 1996).

METHODS

The test item was administered by gavage to three groups, each of twelve male and twelve female Wistar Han:RccHan:WIST strain rats, for approximately six weeks (including a two week pre-pairing phase, pairing, gestation and early lactation for females), at dose levels of 100, 250 and 500 mg/kg bw/day. A control group of twelve males and twelve females was dosed with vehicle alone (Arachis oil BP) over the same treatment period. Two recovery groups, each of five males and five females, were treated with the high dose (500 mg/kg bw/day) or the vehicle alone for forty-two consecutive days and then maintained without treatment for a further fourteen days.

Clinical signs, behavioral assessments, body weight change and food and water consumption were monitored during the study.

Pairing of non-recovery animals within each dose group was undertaken on a one male: one female basis within each treatment group on Day 15 of the study, with females subsequently being allowed to litter and rear their offspring to Day 5 of lactation.

During the lactation phase, daily clinical observations were performed on all surviving offspring, together with litter size and offspring weights and assessment of surface righting reflex. Extensive functional observations were performed on five selected non-recovery males from each dose group after the completion of the pairing phase, and for five selected parental females from each dose group on Day 4 post partum. Urinalysis was performed on five non-recovery males per dose group during the final week of treatment and five non-recovery males and females from each dose group were selected for hematology and blood chemistry assessments prior to termination. Surviving adult non-recovery males were terminated on Day 43 or Day 44, with all females and offspring being killed on Day 5 post partum. Any female which did not produce a pregnancy was terminated on or after Day 25 post coitum. All animals were subjected to a gross necropsy examination and histopathological evaluation of selected tissues was performed. Following forty-two days of treatment, recovery group animals were maintained without treatment for a further fourteen days. Urinalysis was performed on all recovery group males during the final week of the treatment period. In addition, hematological and blood chemical assessments were performed on all recovery group animals at the end of the treatment-free period. These animals were then subjected to a gross necropsy and histopathological examinations of selected tissues was performed.

RESULTS

Mortality: At 100 mg/kg bw/day, one male died during the blood sampling procedure on Day 42; this death was considered to be incidental and unrelated to treatment.

Clinical observations: Treatment at 500 mg/kg bw/day was associated with increased post-dosing salivation in all seventeen male and seventeen female; ten males and eight females also showed noisy respiration during the study. Increased post-dosing salivation was also observed for five males at 100 mg/kg bw/day and the twelve males and eight females at 250 mg/kg bw/day.

Behavioural assessment: There were no treatment-related changes in the behavioural parameters at 100, 250 or 500 mg/kg bw/day.

Functional performance tests: There were no changes in functional performance considered to be related to treatment at 100, 250 or 500 mg/kg bw/day.

Sensory reactivity assessments: There were no inter-group differences in sensory reactivity scores.

Body weight: At 500 mg/kg bw/day, males showed lower body weight gain during the first week of treatment, thereafter there was recovery of body weight gain although body weight gain still tended to be lower than control during the treatment period. Body weight gain of males was slightly superior to control during the recovery period. For females at 500 mg/kg bw/day, lower body weight gain was also observed during the first week of treatment. There was no subsequent effect of treatment on body weight gain during the remainder of the study, including gestation, lactation and recovery phases. At 250 mg/kg bw/day, males showed lower body weight gain during the first week of treatment; body weight gain was considered to be unaffected by treatment for the remaining treatment period and the recovery period. At both sexes at 100 mg/kg bw/day and females at 250 mg/kg bw/day, body weight gain was considered to be unaffected by treatment.

Food consumption: At 500 mg/kg bw/day, food consumption for both sexes was slightly lower than control during the first week of treatment; thereafter food intake was unaffected by treatment for the remainder of the study including gestation, lactation and recovery phases. At 100 and 250 mg/kg bw/day, food consumption for both sexes was considered to be unaffected by treatment throughout the study.

Food consumption efficiency: At 500 mg/kg bw/day, food conversion efficiency for both sexes was slightly lower than control during the first week of treatment. At 100 and 250 mg/kg bw/day, food conversion efficiency for both sexes was considered to be unaffected by treatment.