N,N'-diphenyl-p-phenylenediamine

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

In a screening study for reproductive / developmental toxicity (Matsumoto et al., 2013) according to OECD TG 421, DPPD was applied to rats at 8, 50 and 300 mg/kg bw/d by gavage. Gestation times were significantly elongated in females in the 50 and 300 mg/kg group. From gestation day 22 onwards, severe clinical signs (pale skin, piloerection, hypothermia), mortality, dystocia were observed in females in the 300 mg/kg group. Two females from that group were sacrificed on day 23 of pregnancy, and the following gross pathological findings were observed in these animals: haemorrhage in the lumen of the uterus, incomplete retention and red colour in the lung, and dark red medulla and hardness on the kidney in both animals; hydrothorax in the thoracic cavity, attachment of red content in mucosa of the glandular stomach and recessed area, or red spots in the duodenum in either animal.

The NOAEL with regard to maternal toxicity was 50 mg/kg in this study (based on mortality and clinical signs in the highest dose group), and the NOAEL with regard to reproductive toxicity was 8 mg/kg (elongated gestation lengths at higher doses).

The observed maternal toxicity can be attributed to the significant fertility effect in females, namely prolonged gestation periods. Mechanistic data (see "mode of action analysis / human relevance framework") strongly indicate that DPPD impairs the onset of parturition by interfering with prostaglandins that are critical in this process. As a consequence of a delayed onset of parturition, severe maternal toxicity including mortality can be observed at the end of the gestation period. The association of maternal toxicity with the process of parturition is supported by the observation that no significant effects were found in the 28 -day study by Matsumoto et al. up to the limit dose of 1000 mg/kg. It is further corroborated by the fact that no clinical signs whatsoever were observed in pregnant females until day 22, when the animals should normally start to give birth.

In an older one-generation study submitted to US EPA (OTS0556010, 1993), rats were treated with 300 or 1000 ppm DPPD in feed. Maternal mortality, histopathological changes of the urinary and hepatobiliary tract, decreased numbers of live births and prolonged gestation periods were reported. The LOAEL in this study is 300 ppm for maternal toxicity, reproductive performance and developmental effects. However, the reliability of the study is questionable as it was performed pre-GLP, no guideline was followed, and pregnancies was determined based on the weight of rats. Also, different numbers of pregnant females were used in the different groups. The study was evaluated as a supporting study only.

Effects on developmental toxicity

Description of key information

In the screening study for reproductive / developmental toxicity by Matsumoto et al. (2013), several offspring parameters were dose-dependently decreased, reaching an adverse level in the 300 mg/kg group: no. of pups born, delivery index, no. of live pups, birth index and live birth index. None of these changes was statistically significant.

The above-mentioned alterations in developmental parameters were not very strong and reached an adverse stage only at 300 mg/kg, where severe maternal toxicity including mortality was observed. The developmental effects are therefore considered to be associated with and secondary to these maternal effects. They are not considered to represent specific developmental toxicity.

Mode of Action Analysis / Human Relevance Framework

A mechanism of the prolonged gestation period caused by DPPD has been investigated in the literature:

Marois (Bull. Acad. Natl. Med;1982(3):581-95, 1998) found that a single application of PGF2α on the 21st day of gestation re-establishes the normal onset of parturition in rats that had previously been treated with DPPD from day 14 of gestation onwards. Fujita et al. (Experentia 38(12), 1472, 1982) found that DPPD inhibited the synthesis of PGE in rabbit kidney medulla slices. Yohko et al. (J. Pharm. Pharmacol, 1984, 36:195-197) showed that this inhibition could be reversed by addition of arachidonic acid.

These results indicate that prolongation of the gestation period of DPPD is mediated by an inhibition of PGF2α and/or PGE2.

PGF2α and PGE2 are known to be critical modulators of parturition in rats and mice by induction of luteolysis (PGF2α) and cervix ripening (PGE2), respectively, prior to birth. While the effects in mediated by PGE2 in this context are comparable in rats and humans, a significant interspecies difference exists regarding the influence of PGF2α: In humans and guinea pigs, luteolysis is not a key event to initiate parturition. Birth can begin without a prior decrease in progesterone levels. For mice and rats, this process is, however, critical (Welsh et al., J. Physiol. 569.3(2005) 903-912).

In conclusion, human relevance of this mode of action can thus not be excluded: The DPPD effects mediated by interference with PGE2 may in principle also be relevant to humans; however, interference with PGF2α represents a second crucial modulator in rats, which is irrelevant in humans. It is therefore likely that the rat represents a quantitative and qualitative worst case compared to humans.

Justification for classification or non-classification

Based on the findings from the screening study for reproductive / developmental toxicity by Matsumoto et al., 2013, according to OECD TG 421, a classification into Repr. Cat. 2 (H361f). Further information is expected from a proposed extended one-generation study.