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Key value for chemical safety assessment

Effects on fertility

Effect on fertility: via oral route
Dose descriptor:
NOAEL
70 mg/kg bw/day
Additional information

In a 2-generation GLP reproductive toxicity study, male and female Sprague-Dawley rats were orally administered ptBP via the diet at concentrations of 0 (control), 800 (low-dose), 2500 (mid-dose), or 7500 (high-dose) ppm (Clubb and Jardine, 2006), corresponding to approximately 0, 70, 200, and 600 mg/kg body weight/day. F0 animals were treated for 10 weeks prior to mating, during the mating period, and until termination at weaning of the F1 generation. F1 animals were exposed to ptBP from conception until 11 weeks after weaning, then throughout the mating period and until termination at weaning of the F2 generation. The F2 generation was exposed to ptBP from conception until termination at weaning. No treatment-related clinical signs were observed. Notably, decreases in body weight gain and food consumption were observed in all generations, including during gestation at the mid- and high-dose levels. Reduced pup weights in the F1 and F2 generations (mid- and high-dose groups) and increased pup mortality in the F1 generation (high-dose group) also were observed, likely due to decreased maternal weight gain and food consumption. Several organ weight changes were observed in the F1 and F2 generations at the mid- and high-dose levels. There were no effects on sperm characteristics, estrous cycles, mating performance, fertility, and duration of gestation. Necropsy revealed no obvious effect of treatment on the F0 and F1 adults or on the pups of the F1 and F2 generations; however, key histopathological findings were noted for the ovary in high-dose F1 females and for vagina in mid- and high-dose F0 and high-dose F1 females. Based on the changes observed in this study, the investigators concluded that the reproductive no observed effect level (NOEL) for ptBP was 800 ppm (Clubb and Jardine, 2006).

A range finding study conducted prior to this study examined dose levels of 5000, 10000, and 20000 ppm administered via the diet to male and female F0 generation rats for 2 weeks prior to mating and continuously thereafter through to weaning of the F1 generation animals (Clubb and Jardine, 2006). Parental effects were evident at all dose levels. The animals, particularly at 10000 and 20000 ppm, had reduced food consumption but it was not possible to state whether or not this reduction was due to palatability. The reddened mucosa in the stomach and distended intestines may have been indicative of an irritant effect of the material. A lower pregnancy rate was observed at 20000 ppm, lower number of implants at 10000 and 20000 ppm, and lower pup weights at all levels. Levels of 10000 and 20000 ppm were not considered suitable for use in the subsequent study, and following discussion with the Sponsor, and in the light of the comments from the regulatory authorities, a level of 7500 ppm was agreed for use as the high dose on the subsequent study (described above).

A combined repeated dose and reproductive/developmental toxicity screening test (OECD 422) conducted according to GLP also has been performed in which groups of male and female Sprague-Dawley rats (13/sex/dose) received daily doses of ptBP via oral gavage at doses of 0, 20, 60, and 200 mg/kg body weight/day (Ministry of Health & Welfare, Japan, 1996). Males were administered ptBP over a 6-week period beginning from 14 days prior to mating to 14 days after mating, and females were administered ptBP from 14 days prior to mating to day 4 of lactation.  As vehicle a 0.5% methyl cellulose solution was used. At 200 mg/kg body weight/day one female from the F0 generation was found dead two days after delivery. Five of the 16 newborns from this dam were found dead on the delivery day. The other 11 newborns were dead on the first day of delivery. Gross necropsy of the dam showed sub involution and change in colour (red or black) in the lungs. Histopathologic examination revealed congestion in the lungs; however, this was considered to be an administration mistake. Furthermore, at 200 mg/kg body weight/day some females showed stridor associated with dyspnea, likely caused by irritation of the respiratory tract. This may be related to a secondary effect due to gavage application of an irritating material. There was no significant difference in the number of corpora lutea, number of implantation sites, in the number of pups born, delivery index, number of pups alive, birth index, and live birth index between the control animals and the exposed animals. The plasma concentration of albumin in the males was slightly decreased, accompanied by decrease in plasma protein. A significant decrease in red blood cells and white blood cells in males in the 200 mg/kg body weight/day dose group also was reported. No compound related morphological changes were observed during pathological examination of parental animals. There were no treatment related toxic effects in pregnant and lactating females other than respiratory irritation. The NOAEL for systemic toxicity in the parental generation was considered to be 60 mg/kg body weight/day while the NOAEL for effects on fertility was 200 mg/kg bw/day (MHW, 1996).

Short description of key information:
A 2-generation GLP study in Sprague-Dawley rats in which ptBP was administered in the diet determined a no-observed-effect-level of 800 ppm (approximately 70 mg/kg body weight/day), based on reduced pup weights at 2500 and 7500 ppm in both generations, and increased pup mortality in F1 pups at 7500 ppm. There were no effects on sperm characteristics, estrous cycles, mating performance, fertility, and duration of gestation. An additional GLP study (combined repeated dose and reproductive/developmental toxicity screening study) conducted in Sprague-Dawley rats established a no-observed-adverse-effect level for fertility of 200 mg/kg body weight/day.

Effects on developmental toxicity

Description of key information
Data from a developmental toxicity study conducted according to OECD test guideline 414 are not available.  However, data from a 2-generation GLP study in Sprague-Dawley rats in which ptBP was administered in the diet determined a no-observed-effect-level of 800 ppm (approximately 70 mg/kg body weight/day), based on reduced pup weights at 2500 and 7500 ppm in both generations, and increased pup mortality in F1 pups at 7500 ppm.  An additional GLP study (combined repeated dose and reproductive/developmental toxicity screening study) conducted in Sprague-Dawley rats established a no-observed-adverse-effect level for developmental toxicity of 200 mg/kg body weight/day.
Effect on developmental toxicity: via oral route
Dose descriptor:
NOAEL
70 mg/kg bw/day
Additional information

In a 2-generation GLP reproductive toxicity study, male and female Sprague-Dawley rats were orally administered ptBP via the diet at concentrations of 0 (control), 800 (low-dose), 2500 (mid-dose), or 7500 (high-dose) ppm (Clubb and Jardine, 2006), corresponding to approximately 0, 70, 200, and 600 mg/kg body weight/day. F0 animals were treated for 10 weeks prior to mating, during the mating period, and until termination at weaning of the F1 generation. F1 animals were exposed to ptBP from conception until 11 weeks after weaning, then throughout the mating period and until termination at weaning of the F2 generation. The F2 generation was exposed to ptBP from conception until termination at weaning. No treatment-related clinical signs were observed. In the F0 and F1 generations there were a lower number of implant sites and live pups born at 7500 ppm. The lower numbers of implants may be associated with the lower body weights. At 7500 ppm, in both generations, litter size was slightly smaller than Controls and pup weight and litter weight were also lower than Controls from Day 1 of lactation. At 2500 ppm, in both generations, pup weight and litter weight gain were also lower from Day 14 of lactation. Among F1 pups, weight gain and survival of the pups was lower than that of the Controls suggesting a poor maternal performance. This may have been due, in part at least, to the decreased weight gain/ food consumption performance seen in the F0 animals. For the F2 pups, the survival of the smaller litters was good and exceeded the performance of the Control litters, although the weight gain of the pups over lactation was lower.

There were no effects of treatment on ano-genital distance, or on nipple/areolar retention. At 7500 ppm delays in vaginal opening and preputial separation (occurred 3 and 4 days later than Controls, respectively) were considered to be due to the lower body weight.

At 7500 ppm kidney weight in males was higher than the Controls following covariance analysis in both generations, although the biological significance of this finding is equivocal. At 7500 ppm decreases in females in adrenal gland, ovary and pituitary gland weights achieved statistical significance in both generations. Whilst there was no associated alteration in the normal structure of these organs, as seen by microscopy, it was considered, because of the reproducibility of the organ weight findings in both generations and because these organs are associated with reproductive function, that these findings could not be entirely dismissed.

In the pups, marginal reduction in thymus weight were evident in F1 females at 2500 and 7500 ppm following covariance analysis; spleen weight in F2 males was significantly but slightly decreased. There were no necropsy findings considered to reflect an obvious effect of treatment on either the F0 or F1 adults, or on the weanlings of either generation. In the ovary there was a decrease in growing follicles in F1 females treated at 7500 ppm, which was not clearly evident in the F0 generation. In the F1 animals, the incidence of vaginal epithelium atrophy was increased in severity and incidence at 7500 ppm, but was not evident at 2500 ppm.

Therefore the primary findings from this study were those noted in the ovary and vagina, but these findings do not support each other, as there is no clear consistency in the incidence or severity of the findings: although at 7500 ppm the vagina and ovaries appeared to be more affected in the F1 females, at 2500 ppm the vagina was more affected in the F0 generation.

In conclusion, the only effects of treatment on reproductive performance were reduced pup weights at 2500 and 7500 ppm in both generations, and increased pup mortality in F1 pups at 7500 ppm; many of the findings reported appear to be related to maternal size or performance and/or direct toxicity. The other key findings from this study are those noted in the ovary and vagina. It was therefore considered that the no observed effect level (NOEL) was 800 ppm (Clubb and Jardine, 2006).

A combined repeated dose and reproductive/developmental toxicity screening test (OECD 422) conducted according to GLP also has been performed in which groups of male and female Sprague-Dawley rats (13/sex/dose) received daily doses of ptBPviaoral gavage at doses of 0, 20, 60, and 200 mg/kg body weight/day (Ministry of Health & Welfare, Japan, 1996). Males were administered ptBP over a 6-week period beginning from 14 days prior to mating to 14 days after mating, and females were administered ptBP from 14 days prior to mating to day 4 of lactation. Examination of body weights and gross morphology of the offspring revealed no effects of ptBP, and there were no significant differences in the viability index on day 4 of lactation between the control animals and the exposed animals. However, at 200 mg/kg body weight/day one female from the F0 generation was found dead two days after delivery. Five of the 16 newborns from this dam were found dead on the delivery day. The other 11 newborns were dead on the first day of delivery. There were no morphological abnormalities in the newborns that were dead on the day of delivery. Furthermore, no abnormalities were found in the newborns that underwent necropsy on day 4 of post partum. Gross necropsy of the dam showed sub involution and change in colour (red or black) in the lungs. Histopathologic examination revealed congestion in the lungs; however, this was considered to be an administration mistake.

Overall, no treatment related toxic effects on offspring were reported and a NOAEL of 200 mg/kg body weight/day for reproductive/developmental toxicity was identified (MHW, 1996).

Justification for classification or non-classification

The substance is classified as toxic for reproduction (category 2) according to Regulation (EC) NO. 1272/2008 and classified as Cat. 3 (developmental effects) as set out in Directive 67/548/EEC.

Additional information