Amines, C12-18-alkyldimethyl, N-oxides

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

No other studies concerning toxicity to reproduction are available.

Effect on fertility: via oral route

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

100 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

One screening study (K=1) is available.

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

A combined repeat dose toxicity study with the reproduction/developmental toxicity screening test conducted under GLP according to OECD TG 422 is available for C12 -18 AO [Hansen B (2010)]. In this study rats (CD/Crl:CD(SD); 10 animals/sex/group) were dosed by oral gavage with C12 -18 AO at levels of 0, 40, 100 or 200 mg AO/kg bw/day for two weeks prior to mating,during mating and approximately two weeks after mating (males) and for two weeks prior to mating and continuing up to and including day 3 post-partum or the day prior to sacrifice for females. General toxicity effects in parent animals are discussed in the repeated dose toxicity section. No test item related influence was noted on the female fertility index or pre-coital time at any of the tested dose levels. There were no test item related differences in the number of corpora lutea, implantation sites, in the number and sex of pups, runts or malformed pups. No test item related influence was noted in the values calculated for the gestation length, the gestation index, the birth index and the live birth index between the control group and animals in any test group. No test item related increase in pre-implantation loss was noted in the dams after treatment with 40 or 100 mg AO/kg bw/day throughout the study up to day 3 post-partum. Treatment with 200 mg AO/kg bw/day resulted in a statistically significant (p<= 0.05) increase of the pre-implantation loss by 20.7 % compared to the control (10.5 %). The post implantation loss was not influenced at any dose level. In pups no test item related effects were noted for mortality, abnormal behaviour, mean and total litter weight or external abnormalities at any of the doses tested. Due to the pre-implantation loss noted at the high dose of 200 mg AO/kg bw/day the NOEL for reproduction/developmental toxicity was 100 mg AO/kg bw/day.

In a combined repeat dose toxicity study with the reproduction/developmental toxicity screening test conducted under GLP according to OECD TG 422 C12 -14 AO was administered via gavage to 10 animals/sex/group at 0, 40, 100 or 250 mg/kg bw/day. Males were dosed for at least 14 days prior to mating and during 14 days pairing and female rats for 14 days prior to pairing, through the pairing and gestation period until the offspring reached Day 4 post partum. General toxicity effects on parent animals are discussed in the section on repeat dose toxicity. Microscopic examination of the reproductive organs of males and females treated at 250 mg AO/kg bw/day failed to reveal any abnormalities. Litter size and mean number of pups at first litter check were not affected by treatment. The sex ratio was also not affected. No abnormal pup was noted at any dose level. At 250 mg AO/kg bw/day a statistically significant increase in pups death was observed on postnatal days 0-4. Although the higher postnatal loss was in the range of historical control data, this resulted in a reduced number of pups. Mean pup weight development was reduced at 250 mg AO/kg bw/day. At necropsy, there were no findings in pups. The NOEL for reproduction/developmental toxicity was considered to be 100 mg/kg bw/day.

Short description of key information:
In a combined repeat dose toxicity study with the reproduction/developmental toxicity screening test conducted under GLP according to OECD TG 422 C12-18 AO was administered via gavage to rats. The NOEL for reproduction/developmental toxicity was considered to be 100 mg/kg bw/day.
In a similar study C12-14 AO was administered via gavage to 10 animals/sex/group at 0, 40, 100 or 250 mg/kg bw/day. The NOEL for reproduction/developmental toxicity was considered to be 100 mg/kg bw/day.

Justification for selection of Effect on fertility via oral route:
Only study available

Effects on developmental toxicity

Description of key information

In a prenatal developmental toxicity study in which pregnant female rats were dosed by gavage with C12-14 Ao at 0, 25, 100 or 200 mg AO/kg bw on gestation days 6-19, the NOAEL for both maternal toxicity and developmental toxicity is 25 mg AO/kg/day.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

25 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

There is one prenatal developmental toxicity study (K=1) and one screening study (K=1) available.

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

The key study is a prenatal developmental toxicity study performed under GLP according to EPA OTS 798.4900 [York RG (1999). In this study presumed pregnant female rats (Crl:CDBR VAF/Plus; 25 animals/group) were dosed by oral gavage with C12 -14 AO at 0, 25, 100 or 200 mg AO/ kg bw daily on days 6 through 19 of presumed gestation (DG 6-19). The rats were observed for viability at least twice a day and for general appearance weekly during acclimation and on DG 0. The rats were also examined for clinical observations of effects of the test substance, abortions, premature deliveries and deaths immediately before and approximately 60 minutes after dosage and on the day of sacrifice (DG 20). Body weights were recorded weekly during acclimation, on DG 0, daily during the dosage period and on DG 20. Feed consumption values were recorded on DGs 0, 6, 9, 12, 15, 18, and 20.

All surviving rats were sacrificed on DG 20, and a gross necropsy of the thoracic, abdominal and pelvic viscera was performed. Caesarean-sectioning and subsequent fetal observations were conducted without knowledge of dosage group in order to minimize bias. The number of corpora lutea in each ovary was recorded. The uterus of each rat was excised and examined for pregnancy, number and distribution of implantations, live and dead fetuses and early and late resorptions. The gravid uterus was weighed. Each fetus was removed from the uterus and subsequently weighed and examined for sex and gross external alterations. Approximately one-half of the fetuses in each litter were examined for soft tissue alterations using a variation of the microdissection techniques. The remaining fetuses in each litter were examined for skeletal alterations.

Two rats in the 200 mg AO/kg/day dosage group died; one of these deaths was attributed to the test substance, the other was the result of an intubation error. All other rats survived until scheduled sacrifice. Excessive salivation, rales, urine-stained abdominal fur, brown or red perioral substance, labored breathing and gasping occurred in 100 mg AO/kg/day dosage group rats and in significantly increased numbers of 200 mg AO/kg/day dosage group rats. Additionally, chromorhinorrhea occurred in one and two rats in these two dosage groups, respectively. Observations of brown or red perivaginal substance, emaciation, brown perianal or perinasal substance, dehydration, ungroomed coat and soft or liquid feces occurred in one or two rats in the 200 mg AO/kg/day dosage group. All necropsy observations were considered unrelated to the test substance.

Rats in the 100 and 200 mg AO/kg/day dosage groups had significantly reduced body weight gains for the entire dosage period (calculated as days 6 to 20 of gestation) and body weight gains were significantly reduced in the 200 mg AO/kg/day dosage group for the entire gestation period (DGs 0 to 20). Body weights were significantly reduced in the 200 mg AO/kg/day dosage group on DGs 11 through 20. The gravid uterine weight and the corrected DG 20 maternal body weight (DG 20 body weight minus the gravid uterine weight) were significantly reduced in the 200 mg AO/kg/day dosage group.

Absolute (g/day) and relative (g/kg/day) feed consumption values for the entire dosage period (calculated as DGs 6 to 20) and the entire gestation period (DGs 0 to 20) were significantly reduced in the 200 mg AO/kg/day dosage group. Relative feed consumption values for the entire dosage period were also significantly reduced in the 100 mg AO/kg/day dosage group, while values for the gestation period were significantly reduced in the 25, 100 and 200 mg AO/kg/day dosage groups.

Male and female fetal body weights were significantly reduced in the 200 mg AO/kg/day dosage group. Live litter size was decreased and the number of early resorptions was increased in the 200 mg AO/kg/day dosage group, but apparently as the result of one dam in this dosage group that had a litter consisting of 16 early resorptions.

The percentages of fetuses and litters with alterations in the 200 mg AO/kg/day dosage group were significantly increased and reflected delays in skeletal ossification related to the significantly reduced fetal body weights in this group. These delays in ossification included significant increases in the fetal and/or litter incidences of bifid thoracic vertebrae centra, incompletely and/or not ossified 1st or 2nd sternal centra, incompletely ossified pubes and significant decreases in the numbers of ossified caudal vertebrae, sternal centra and metacarpals. Additionally, delays in ossification occurred in the 100 mg AO/kg/day dosage group and included a significant increase in the litter incidence of bifid thoracic vertebrae centra.

NOAEL for both maternal toxicity and developmental toxicity is 25 mg AO/kg/day.

A combined repeat dose toxicity study with the reproduction/developmental toxicity screening test conducted under GLP according to OECD TG 422 is available for C12 -18 AO [Hansen B (2010)]. In this study rats (CD/Crl:CD(SD); 10 animals/sex/group) were dosed by oral gavage with C12 -18 AO at levels of 0, 40, 100 or 200 mg AO/kg bw/day for two weeks prior to mating, during mating and approximately two weeks after mating (males) and for two weeks prior to mating and continuing up to and including day 3 post-partum or the day prior to sacrifice for females. General toxicity effects in parent animals are discussed in the repeated dose toxicity section. No test item related influence was noted on the female fertility index or pre-coital time at any of the tested dose levels. There were no test item related differences in the number of corpora lutea, implantation sites, in the number and sex of pups, runts or malformed pups. No test item related influence was noted in the values calculated for the gestation length, the gestation index, the birth index and the live birth index between the control group and animals in any test group. No test item related increase in pre-implantation loss was noted in the dams after treatment with 40 or 100 mg AO/kg bw/day throughout the study up to day 3 post-partum. Treatment with 200 mg AO/kg bw/day resulted in a statistically significant (p<= 0.05) increase of the pre-implantation loss by 20.7 % compared to the control (10.5 %). The post implantation loss was not influenced at any dose level. In pups no test item related effects were noted for mortality, abnormal behaviour, mean and total litter weight or external abnormalities at any of the doses tested. Due to the pre-implantation loss noted at the high dose of 200 mg AO/kg bw/day the NOEL for reproduction/developmental toxicity was 100 mg AO/kg bw/day.

In a combined repeat dose toxicity study with the reproduction/developmental toxicity screening test conducted under GLP according to OECD TG 422 C12 -14 AO was administered via gavage to 10 animals/sex/group at 0, 40, 100 or 250 mg/kg bw/day. Males were dosed for at least 14 days prior to mating and during 14 days pairing and female rats for 14 days prior to pairing, through the pairing and gestation period until the offspring reached Day 4 post partum. Litter size and mean number of pups at first litter check were not affected by treatment. The sex ratio was also not affected. No abnormal pup was noted at any dose level. At 250 mg AO/kg bw/day a statistically significant increase in pups death was observed on postnatal days 0-4. Although the higher postnatal loss was in the range of historical control data, this resulted in a reduced number of pups. Mean pup weight development was reduced at 250 mg AO/kg bw/day. At necropsy, there were no findings in pups. The NOEL for reproduction/developmental toxicity was considered to be 100 mg/kg bw/day.

No data from prenatal developmental toxicity studies in a second species, typically the rabbit, are available for the amine oxides.

As discussed in the toxicokinetics section, amine oxides are extensively metabolised after oral administration as shown by at least 10 metabolites characterised in urine. In addition a substantial proportion of the dose is completely bio transformed to CO₂and therefore eliminated by exhalation. There were several different pathways of metabolism proposed:

·        Oxidative degradation of the alkyl side chain by ω-oxidation to form a carboxylic acid followed by the loss of two-carbon fragments by sequential β-oxidation. This route of metabolism is frequently observed in similar substances to the test substance.

·        Hydroxylation of the alkyl side chain at a position four or five carbons from the end of the chain.

·        Reduction of the amine oxide group to the parent amine.

The metabolism of amine oxides is considered to be likely to be similar between different mammalian species including the rabbit and the rat.

A review of the literature has shown that some developmental toxicity data are available for amines and it is considered appropriate to read across to this class of chemicals as they are a relevant intermediate in the metabolic pathway of amine oxides.

In a study performed using cis-9-octadecenylamine (CAS No. 112-90-3) New Zealand White rabbits (22 artificially-inseminated females/group) were administered the test substance in corn oil via gavage at doses of 0, 3, 10 or 30 mg/kg bw/day for days 6 through 18 of gestation [US EPA (2010) ]. Two females died in the high-dose group. Clinical signs (irritation of the lips and chin, laboured breathing and rales) of toxicity were observed at 10 and 30 mg/kg bw/day. Dose-dependent body weight losses or reduced gains and reduced food consumption occurred at the 10 and 30 mg/kg bw/day (significant only in the high dose group). There were no local effects on the gastrointestinal tract of the does. There were no treatment-related effects on the foetuses examined. The NOAEL (maternal toxicity) was 3 mg/kg bw/day and the NOAEL (developmental toxicity) was 30 mg/kg bw/day (highest dose tested).

In a study performed using N-methyl-N-octadecyl-1-octadecanamine (CAS No. 4088-22-6) pregnant New Zealand White rabbits (16/dose) were administered the test substance in corn oil via gavage at doses of 0, 50, 250 or 1000 mg/kg bw/day for days 6 through 18 of gestation [US EPA (2010)]. No treatment-related effects were found on pregnancy rate, mortality, abortions or gestation length. There was a high post implantation loss seen in the high dose group but this was not significant given the unusually high loss also observed in controls. No treatment-related effects were found on foetal size, foetal sex or mortalities; however slight reductions in foetal weight at 250 and 1000 mg/kg bw/day were observed and possible embryolethality at 1000 mg/kg bw/day cannot be disregarded. The NOAEL (maternal toxicity) was 50 mg/kg bw/day and the NOAEL (developmental toxicity) was 250 mg/kg bw/day.

The absence of developmental toxicity effects at dose levels below those at which maternal systemic toxicity was observed in studies performed in rats using C12-18 AO and C12-14 AO and in rabbits using cis-9-octadecenylamine or N-methyl-N-octadecyl-1-octadecanamine (surrogates for the metabolic intermediates of amine oxides) indicates that there is no concern for developmental toxicity from amine oxides.

Toxicity to reproduction: other studies

Additional information

No other studies concerning toxicity to reproduction are available.

Justification for classification or non-classification

In the prenatal developmental toxicity study performed using C12-14 AO effects were seen in pups from 100 mg/kg bw/day. These effects included delayed ossification and reduced body weight (significant at 200 mg/kg bw/day). In addition, live litter size was decreased and the number of early resorptions increased in the 200 mg/kg bw/day group (results skewed by one dam with a high level of resorptions). These effects were seen at levels where maternal toxicity was evident. The maternal effects noted included excessive salivation, rales, urine-stained abdominal fur, brown or red perioral substance, laboured breathing and gasping starting from 100 mg/kg bw/day and significant at 200 mg/kg bw/day, significantly reduced body weight gain in both groups and significantly reduced body weight at 200 mg/kg bw/day and mortality (one female) at 200 mg/kg bw/day. On this basis it is considered that the effects seen in pups at the high dose are asecondary non-specific consequence of maternal toxicity.

In the OECD TG 422 study performed using C12-14 AO total locomotor activity in females was statistically significantly reduced at 250 mg/kg bw/day. At 100 mg/kg bw/day lesions in the forestomach were noted at necropsy and histopathology. There were no effects on litter size, mean number of pups at first litter check, sex ratio or pup abnormality. At 250 mg AO/kg bw/day there was a statistically significant increase in pup death on postnatal days 0-4. The higher postnatal loss was in the range of historical controls. Mean pup weight development was reduced at 250 mg AO/kg bw/day. At necropsy there were no findings in pups. The NOEL for reproductive/developmental toxicity was considered to be 100 mg/kg bw/day in this study. The NOAEL for maternal toxicity was established at 40 mg AO/kg/day. In this study it is considered that the effects seen in pups at the high dose are a secondary non-specific consequence of maternal toxicity.

In the OECD TG 422 study performed using C12-18 AO there were no effects on female fertility index or pre-coital time at any dose level. There were no differences inthe number of corpora lutea, implantation sites, in the number and sex of pups, runts or malformed pups. There were no differences in gestation length, the gestation index, the birth index and the live birth index between the control group and animals in any test group. Treatment with 200 mg AO/kg bw/day resulted in a statistically significant increase of the pre-implantation loss compared to the control. The post implantation loss was not influenced at any dose level. In pups there were no differences in mortality, abnormal behaviour, mean and total litter weight or external abnormalities at any of the doses tested.

The NOEL for reproductive/developmental toxicity was considered to be 100 mg/kg bw/day in this study. The NOAEL for maternal toxicity was established at 40 mg AO/kg/day. In this study it is considered that the effects seen in pups at the high dose are a secondary non-specific consequence of maternal toxicity.

Based on the findings in these studies it is concluded that any effects seen in pups are a secondary non-specific consequence of maternal toxicity and therefore no classification for reproductive toxicity is warranted.