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Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information
No further data available
Effect on fertility: via oral route
Endpoint conclusion:
no study 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

There is no information available from a specific reproduction toxicity study on Etherdiamine C13i/acetate.

A 28-day study on Etherdiamine C13i/acetate included the evaluation of reproductive organs. The following organs were weighted and microscopically examined: Epididymides, prostate and seminal vesicles incl. coagulating glands, testis, ovaries and uterus. Dose levels were 0, 3, 9, and 25 mg/kg bw/day dosed by gavage. In the high dose group, 2/5 males and 1/5 males from the recovery group showed reduced size seminal vesicles upon gross examination, and of prostate gland in one of these high dose group animals. Organ weights showed a dose dependent reduced weight, which was significant in the high dose males for prostate (with seminal vesicles incl. coagulating glands) (44%) and Epididymides (18%). Only for the high dose group for prostate effects remained when based on body weight ratio’s. The recovery groups showed no differences for these organ weights, indicating that observed effects are reversible and likely secondary to the general weight loss. The organ weights for ovaries and uterus showed no significant changes in females. Histopathology only indicated minimal reduced secretion in the prostate gland and minimal reduced colloid in the seminal vesicles in high dose group males. Only minimal reduced colloid in the seminal vesicles was still present in one recovery male.

Considering that the lowest dose of 3 mg/kg bw represented a LOAEL rather than a NOAEL, the effects on prostate and seminal vesicles in the high dosed males at 25 mg/kg which was resolved after a 14-day recovery period are not indicative for a specific reproductive organ toxicity.

In a 90-days study Wistar rats were administered 0, 0.5, 2.2 or 8.8 mg/kg bw/d of Etherdiamine C13i/acetate. Examinations were performed according to OECD 408 with additionally oestrous cycle determination. There were no indications of possible reproductive toxicity based on the parameters determined in this study. Estrous cycle length/regularity appeared unaffected during the period in which estrous cycle length was determined (Day 76-91), and histopathological examination of the male and female reproductive organs did not show lesions that were directly related to treatment.

Inactive uterus (cervix) epithelium was observed in 4/10 females at 2 mg/kg. In general, this finding is probably related to poor health condition. Atrophy/mucification of the vagina epithelium was seen in 1/1 females at 0.5 mg/kg and in 1/10 females a 2 mg/kg.

Also in this study, the lowest dose of 0.5 mg/kg represented a LOAEL rather than a NOAEL, and the effects observed on the uterus epithelium (‘inactive uterus’) observed at 2 mg/kg are not indicative for a specific reproductive organ toxicity.

Also the study ofEtherdiamine C13i/acetate in a prenatal developmental toxicity study in rats has not shown reproductive effects.

Literature does not provide additional information. Comparable fatty nitrile derivatives (like diamines) have not shown a concern for reproduction toxicity.

The likelihood of exposures to Etherdiamine C13i/acetate is low considering use and physical properties: The substance is very corrosive and its use is limited to industrial settings under controlled conditions. Also the high boiling point (> 300 °C) and low vapour pressure (0.005 Pa at 25°C) indicate that the potential for inhalation is not significant to justify this study.

 

In view of limited exposures, lack of effects on reproductive organs observed in repeated dose studies in contrast to severe systemic toxicity observed in these studies, and lack of effects on reproduction seen in reproduction and developmental toxicity studies on comparable fatty amine-like substances, there are no concerns for reproduction toxic effects by Etherdiamine C13i/acetate that would warrant further studies for reproduction toxicity.


Short description of key information:
Available repeated dose studies on Etherdiamine C13i/acetate do not indicate adverse effects on reproductive organs, including oestrous cycle evaluation and histopathological examination of the male and female reproductive organs. Prenatal developmental toxicity study on Etherdiamine C13i/acetate showed no indications reproductive effects. Also comparable fatty amine-like substances lack of effects on reproduction.

Justification for selection of Effect on fertility via oral route:
Likelihood of exposures to Etherdiamine C13i/acetate is very low with use limited to industrial settings under controlled conditions. Available repeated dose studies do not indicate adverse effects on reproductive organs, including oestrous cycle evaluation and histopathological examination of the male and female reproductive organs, and the prenatal developmental toxicity study on Etherdiamine C13i/acetate showed no reproductive effects. Also comparable fatty amine-like substances lack effects on reproduction.

Justification for selection of Effect on fertility via inhalation route:
Likelihood of exposures via inhalation is low considering use and physical properties: The substance is very corrosive and its use is limited to industrial settings under controlled conditions. Also the high boiling point (> 300 °C) and low vapour pressure (0.005 Pa at 25°C) indicate that the potential for inhalation is not significant to justify this study.

Justification for selection of Effect on fertility via dermal route:
Substance is corrosive. Effects will be characterized by local corrosive effects that are related to duration, quantity and concentration, rather than by systemic toxicity due to dermal uptake. Related to corrosive properties, exposures are likely to be limited in view of protective measures taken.

Effects on developmental toxicity

Description of key information
In an oral OECD 414 prenatal developmental toxicity study in rats with Etherdiamine C13i/acetate, no maternal NOAEL was determined based on observation of granulocytic (necrotizing) inflammation in the ileum and granulomas with or without necrosis in the mesenteric lymph nodes in females starting at the lowest dose of 3 mg/kg. The developmental NOAEL was established at 3 mg/kg bw. No teratogenic effects were observed.
Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Oct-Dec 2014
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.
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
2001
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.31 (Prenatal Developmental Toxicity Study)
Version / remarks:
2008
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
Version / remarks:
1998
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no
Species:
rat
Strain:
other: RccHan:WIST
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Laboratories BV, Venray, The Netherlands
- Sex: females; untreated females were mated at the supplier, and arrived at the test facility at day 0 or 1 post-coitum (day 0 post-coitum was the day of successful mating).
- Age at study initiation: Approximately 10-14 weeks.
- Weight at study initiation: weight range at start of treatment was 179-276 gr.
- Fasting period before study: no
- Housing: individually in Macrolon plastic cages with sterilized sawdust as bedding material and paper as cage-enrichment/nesting material
- 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
- Temperature (°C): 20.5-20.9
- Humidity (%): 42-53
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 16 Nov to 04 Dec 2014
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 test substance and vehicle. No correction was made for the purity/composition of the test substance. Formulations were placed on a magnetic stirrer during dosing.

Storage conditions of formulations: at ambient temperature

Justification for use and choice of vehicle (if other than water): based on trial formulations performed at WIL Research Europe BV 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 during the treatment phase (21 November 2014), according to a validated method (Project 503896). Samples of formulations were analyzed for homogeneity (highest and lowest concentration) and accuracy of preparation (all concentrations). Stability in vehicle over 6 hours at room temperature under normal laboratory light conditions 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. 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
The concentrations analysed in the formulations of Group 2, Group 3 and Group 4 were in agreement with target concentrations (i.e. mean accuracies between 85% and 115%).
A small response at the retention time of the test substance was observed in the chromatograms of the Group 1 formulations. It was considered to derive from carry-over since a similar response was obtained in the analytical blanks.

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

Stability
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:
not relevant
Duration of treatment / exposure:
From days 6 to 20 post-coitum, inclusive.
Frequency of treatment:
Once daily for 7 d/w.
Duration of test:
15 days
Remarks:
Doses / Concentrations:
0, 3, 7 and 15 mg/kg bw/day
Basis:
actual ingested
No. of animals per sex per dose:
22
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale:
Dose levels were based on results of a 14-Day dose range finding study in which 6 females/dose were exposed to 0, 3, 9 and 25 mg/kg bw/d from days 6 to 20 post-coitum inclusive (See results under "any other information").

Maternal examinations:
CAGE SIDE OBSERVATIONS:
- Time schedule: at least twice daily

DETAILED CLINICAL OBSERVATIONS:
- Time schedule: at least once daily from Day 1 post-coitum onwards up to the day prior to necropsy. The time of onset, grade and duration of any observed signs were recorded.

BODY WEIGHT:
- Time schedule for examinations: days 1, 3, 6, 9, 12, 15, 18 and 21 post-coitum.

FOOD CONSUMPTION:
- Time schedule: days 1-3, 3-6, 6-9, 9-12, 12-15, 15-18 and 18-21 post-coitum. Relative food consumption (g/kg bw/d) was also recorded.

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

NECROPSY:
all animals surviving to day 21 post-coitum, all moribund animals and the animal wiht premature delivery were sacrificed and subjected to an external, thoracic and abdominal examination, with special attention being paid to the reproductive organs. During abdominal examination, special attention was paid to the gastrointestinal tract and mesenteric lymph nodes. HIstopathological examination of stomach, duodenum, jejunum, ileum and mesenteric lymph node were executed. Based on previous studies, no histopathological examination was performed on the lower parts of the intestine (caecum, colon and rectum).
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight
- Number of corpora lutea
- Number of implantations
- Number of early and late resorptions
Fetal examinations:
The following parameters were examined in F1 offspring:
number and distribution of live and dead fetuses, fetal weight, sex of each fetus from ano-genital distance and from gonadal inspections, presence of gross abnomalies.

NECROPSY:
External, visceral and skeletal findings were recorded as developmental variations or malformations for all viable fetuses. Visceral findings were also recorded for dead fetuses. Heads were examined from approximately one-half of the fetuses.
Statistics:
- If the variables could be assumed to follow a normal distribution, the Dunnett-test (many-to-one t-test) based on a pooled variance estimate was applied for the comparison of the treated groups and the control group.
- The Steel-test (many-to-one rank test) was applied if the data could not be assumed to follow a normal distribution.
- The Fisher Exact-test was applied to frequency data.
- The Mann Whitney test was used to compare mean litter proportions (percent of litter) of the number of viable and dead fetuses, early and late resorptions, total resorptions, pre- and post-implantation loss, and sex distribution.
- Mean litter proportions (percent per litter) of total fetal malformations and developmental variations (external, visceral and skeletal), and each particular external, visceral and skeletal malformation or variation were subjected to the Kruskal-Wallis nonparametric ANOVA test to determine intergroup differences. If the ANOVA revealed statistically significant (p<0.05) intergroup variance, Dunn’s test was used to compare the compound-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. Test statistics were calculated on the basis of exact values for means and pooled variances. Individual values, means and standard deviations may have been rounded off before printing. Therefore, two groups may display the same printed means for a given parameter, yet display different test statistics values.

No statistics were applied for data on maternal survival, pregnancy status, group mean numbers of dead fetuses, early and late resorptions, and pre- and post-implantation loss.
Indices:
Pre-implantation loss (%) = ((number of corpora lutea - number of implantation sites)/ number of corpora lutea) x 100

Post-implantation loss (%) = ((number of implantation sites - number of live fetuses)/ number of implantation sites) x 100

Viable fetuses affected/litter (%) = (number of viable fetuses affected/litter / number of viable fetuses/litter) x 100
Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
Mortality:
At the highest dose tested (15 mg/kg bw/d) two females were euthanized in extremis on respectively Days 17 and 15 post-coitum. Toxicologically relevant clinical signs included hunched posture, piloerection and a lean appearance. Furthermore, they had reduced food consumption, and a body weight loss of -3% and -10% was recorded for these females. At necropsy, a thickened wall of the ileum was noted for one female. Another female had no macroscopic abnormalities. Both females were pregnant, with 10 live fetuses and 13 normal implantations in development, respectively.
All remaining females survived until scheduled necropsy.

Clinical signs:
Toxicologically relevant clinical signs were seen at 15 mg/kg bw/d only. These included lethargy (1 female), hunched posture (7 females), piloerection (12 females) and lean appearance (3 females).
Salivation seen after dosing at 3, 7 and 15 mg/kg bw/d was considered not toxicologically relevant, considering the nature and minor severity of the effect and its time of occurrence (i.e. after dosing). This sign was considered to be a physiological response related to taste of the test substance rather than a sign of systemic toxicity.

Body weight:
At 15 mg/kg bw/d, body weights were significantly lower than controls from Days 15-21 post coitum. Body weight gains were significantly lower from Days 9-21 post coitum for females at 7 and 15 mg/kg bw/d with a dose-response relationship. Likewise, body weight gain corrected for the gravid uterine weight was significantly lower at 7 mg/kg bw/d as compared to the concurrent control group, and body weight loss (group mean of -4.6%) was observed at 15 mg/kg bw/d.

Food consumption:
Absolute and relative food consumption were significantly lower for females at 7 and 15 mg/kg bw/d than controls during the entire treatment period (not statistically significant for absolute food consumption in the 7 mg/kg bw/d group from Days 15-18 post-coitum) with a dose response relationship.

Macroscopy:
Treatment related macroscopic findings were seen in the gastrointestinal tract and in the mesenteric lymph nodes at all tested dose levels.
Enlarged mesenteric lymph nodes were observed for 2, 8, 10 and 14 females at 0, 3, 7 and 15 mg/kg bw/d, respectively. Furthermore, reddish discolouration of the mesenteric lymph nodes was noted for one female at 15 mg/kg bw/d.
A thickened ileum was seen for 1, 4 and 6 females at 3, 7 and 15 mg/kg bw/d, respectively. A thickened duodenum was noted for 3, 5 and 5 females at 3, 7 and 15 mg/kg bw/d, respectively. In addition, one female at 7 mg/kg bw/d had a dark red focus on her duodenum. One female at 7 mg/kg bw/d and one female at 15 mg/kg bw/d had a thickened jejunum.

Microscopy:
Test substance related adverse microscopic findings were present in the small intestines (duodenum, jejunum, ileum) and mesenteric lymph node of females starting at 3 mg/kg bw/d.
Duodenum:
- (Foamy) macrophages in 2/22 females (1 minimal, 1 slight) at 15 mg/kg bw/d.
- Granulocytic (necrotizing) inflammation in 1/22 female (1 minimal) at 7 mg/kg bw/d and 4/10 females (2 slight, 2 moderate) at 15 mg/kg bw/d.

Jejunum:
- (Foamy) macrophages in 6/22 females (6 minimal) at 15 mg/kg bw/d.
- Granulocytic (necrotizing) inflammation in 2/22 female (2 moderate) at 15 mg/kg bw/d.

Ileum:
- (Foamy) macrophages in 10/22 females at 3 mg/kg bw/d (7 minimal, 3 slight), in 21/22 females at 7 mg/kg bw/d (8 minimal, 13 slight) and in 21/22 females at 15 mg/kg bw/d (7 minimal, 13 slight, 1 moderate).
- Granulocytic (necrotizing) inflammation at increased incidence and/or severity in 9/22 females at 3 mg/kg bw/d (3 minimal, 2 slight, 4 moderate), in 16/22 females at 7 mg/kg bw/d (4 minimal, 8 slight, 4 moderate) and in 21/22 females at 15 mg/kg bw/d (8 minimal, 7 slight, 5 moderate, 1 marked), compared to in 1/22 females at 0 mg/kg bw/d (1 minimal).

Mesenteric lymph node:
- Granulomas with or without necrosis in 8/22 females at 3 mg/kg bw/d (2 minimal, 2 slight, 2 moderate, 2 marked), in 14/22 females at 7 mg/kg bw/d (8 slight, 5 moderate, 1 marked) and in 18/22 females at 15 mg/kg bw/d (1 minimal, 6 slight, 8 moderate, 3 marked).
- Extranodal inflammation with or without peritonitis in 6/22 females at 3 mg/kg bw/d (3 minimal, 1 slight, 2 moderate), in 15/22 females at 7 mg/kg bw/d (3 minimal, 7 slight, 4 moderate, 1 marked) and in 16/22 females at 15 mg/kg bw/d(6 minimal, 6 slight, 4 moderate).
- Increased incidence and/or severity of increased macrophage foci in 9/22 females at 3 mg/kg bw/d (8 minimal, 1 slight), in 10/22 females (10 minimal) at 7 mg/kg bw/d and in 14/22 females (12 minimal, 2 slight) at 15 mg/kg bw/d, compared to in 1/22 females at 0 mg/kg bw/d (1 minimal)
- Increased cellularity of cortex and/or medullary cords was noted in 1/22 females at 3 mg/kg bw/d (1 minimal), in 6/22 females at 7 mg/kg bw/d (6 slight) and in 1/22 females at 15 mg/kg bw/d (1 minimal).

No treatment-related effects were noted on the number of corpora lutea, implantation sites or pre-implantation loss.
One female at 15 mg/kg bw/d had an early delivery of 13 live pups on Day 21 post coitum. The only relevant findings were reduced food consumption from Day 15 post-coitum, reduced body weight gain on Day 18 post-coitum, and enlarged mesenteric lymph nodes at necropsy. Since this early delivery was an isolated finding, it was considered as a chance finding and not treatment related.

Examination of cage debris of pregnant females revealed no signs of abortion or premature birth for the remaining females in this study.

The numbers of pregnant females were 20, 21, 21 and 22 females in the control, 3, 7 and 15 mg/kg bw/d groups, respectively. Two control females, one female at 3 mg/kg bw/d and one female at 7 mg/kg bw/d were non-pregnant. This was not considered treatment related as treatment started from implantation onwards (i.e. Day 6 post-coitum).
Dose descriptor:
NOAEL
Basis for effect level:
other: maternal toxicity
Remarks on result:
not determinable
Remarks:
no NOAEL identified
Dose descriptor:
NOAEL
Effect level:
ca. 3 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: developmental toxicity
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
No treatment-related effects were noted on the number of viable or dead fetuses, early or late resorptions, or post-implantation loss.
Since two high dose females had to be euthanized on Days 15 and 17 post-coitum due to their poor conditions, the number of litters with viable fetuses available for fetal examination was 20, 21, 21 and 20 in respectively the control, 3, 7 and 15 mg/kg bw/d group. The 20 litters in the high dose group includes the litter 77 that delivered on Day 21 post-coitum. Data from this latter litter are given in a separate table with individual data and not included in the summary data tables.
Litter sizes were not affected by treatment up to 15 mg/kg bw/d. The mean number of fetuses per group was 11.6, 11.7, 11.6 and 10.8 in the control, 3, 7 and 15 mg/kg bw/d groups, respectively. The slightly lower mean litter size noted for the high dose group was not considered to be toxicologically relevant as it was still within normal limits.
No treatment-related effects on sex ratio were observed.
Treatment at 15 mg/kg bw/d resulted in significantly lower fetal body weights (both sexes). Mean fetal body weights (sexes combined) were 5.1, 5.1, 5.0 and 4.6 for the control, 3, 7 and 15 mg/kg bw/d groups, respectively.

External examination revealed malformations of the tail for two fetuses in two different litters at 7 mg/kg bw/d. One fetus had a filamentous tail and another had a small tail (1 mm in length). In the absence of comparable findings at the higher dose level of 15 mg/kg bw/d, these isolated findings were not considered to be treatment related. No treatment-related effects on fetal external morphology were observed.

Visceral malformations were noted for two fetuses in two different litters at the lowest dose level tested. These included one fetus with situs inversus and another fetus with external hydrocephaly. At the isolated incidence and in the absence of any visceral malformation in the two higher dose groups, these abnormalities were considered as chance findings and thus not to be related to treatment. No treatment-related visceral findings were observed in any dose group.

A higher incidence of unossified metatarsal #1 was noted for all treated groups. The mean litter proportion was 8.6%, 10.2% and 18.9% in the 3, 7 and 15 mg/kg bw/d groups, respectively, as compared to 2.9% in the control group. None of these changes was statistically significant. It should, however, be noted that the litter proportion for the high dose group was even higher, since the 7 dead fetuses from litter 77 were not included.
There was also a trend towards a higher incidence of unossified sternebae in the 7 and 15 mg/kg bw/d groups. The mean litter proportion of unossified sternebra #5 was 1.6%, 4.5% and 4.6% in the 3, 7 and 15 mg/kg bw/d groups, respectively, as compared to 2.4% in the control group. In addition, unossified sternebra #2 with or without unossified vertebral centra and reduced ossification of the pubis was noted for 3 fetuses from the same litter.
All remaining skeletal variations recorded in this study were not considered to be treatment related as they occurred infrequently, occurred at frequencies that were within the range of available historical control data or were noted in one control fetus only.

There were no treatment related skeletal malformations.
Malformations of the sternebrae (severely malaligned and/or fused sternebrae) were noted for one low dose fetus and two high dose fetuses. Furthermore, there was one low dose fetus with a rib anomaly. At the isolated incidence and in the absence of a dose related response, these skeletal malformations were considered as chance finding rather than to be related to treatment. The finding of bent limb bones for one fetus was by no means related to treatment as this was a fetus of the control group, treated with the vehicle alone.

Abnormalities:
not specified
Developmental effects observed:
not specified
Conclusions:
In an oral OECD 414 prenatal developmental toxicity study in rats with Etherdiamine C13i/acetate, no maternal NOAEL was determined based on observation of granulocytic (necrotizing) inflammation in the ileum and granulomas with or without necrosis in the mesenteric lymph nodes in females starting at the lowest dose of 3 mg/kg. The developmental NOAEL was established at 3 mg/kg bw. No teratogenic effects were observed.
Executive summary:

In a prenatal developmental toxicity study rats were exposed to 0, 3, 7 or 15 mg/kg bw/d of the test substance once daily by oral gavage from days 6 to 20 post-coitum inclusive. The control group received the vehicle propylene glycol. Examinations were according to OECD 414 (2001) with additional histopathology of the stomach, small intestine and mesenteric lymph nodes.

Two high dose females at 15 mg/kg were euthanized moribund on Days 15 and 17 post-coitum, respectively. Toxicologically relevant clinical signs (lethargy, hunched posture, piloerection, and lean appearance) were noted at 15 mg/kg only. Treatment at 7 and 15 mg/kg resulted in reduced body weights, body weight gains, body weight corrected for uterine weight, and food consumption.

Treatment related adverse morphological alterations were present in the small intestine and the draining mesenteric lymph node of animals in all treated groups. At necropsy, a thickened wall of the duodenum, ileum and/or jejunum was noted which at the microscopic level correlated with (foamy) macrophages in the lamina propria and/or granulocytic (necrotizing) inflammation of the duodenum, jejunum and ileum. In general, these changes were dose related with the highest incidence and severity seen in the ileum. Enlarged mesenteric lymph nodes recorded at necropsy correlated with granulomas with or without necrosis (up to marked), extranodal inflammation with or without peritonitis, increased incidence, severity of increased macrophage foci, increased cellularity of cortex and/or medullary cords. Some of these findings were also noted in the 2 high dose females sacrificed moribund after 9 or 12 treatment days.

Based on the morphological data it is most likely that the (necrotizing) inflammation in the intestine caused by exposure to the test substance spreads to the draining mesenteric lymph nodes with possible breakthrough of granulomas into the abdominal cavity. Treatment at 15 mg/kg resulted in significantly lower fetal body weights (both sexes).This is most likely related to the reduced food consumption and body weight loss seen in the mothers.

A higher incidence of unossified metatarsal #1 was noted for all treated groups. The mean litter proportion was 8.6%, 10.2% and 18.9% in the 3, 7 and 15 mg/kg groups, respectively, as compared to 2.9% in the control group. None of these changes was statistically significant. It should, however, be noted that the litter proportion for the high dose group was in actuality even higher, since the 7 dead fetuses from litter 77 were not included in the group mean count.

There was also a trend towards a higher incidence of unossified sternebae in the 7 and 15 mg/kg groups. The mean litter proportion of unossified sternebra #5 was 1.6%, 4.5% and 4.6% in the 3, 7 and 15 mg/kg groups, respectively, as compared to 2.4% in the control group. In addition, unossified sternebra #2 with or without unossified vertebral centra and reduced ossification of the pubis was noted for 3 fetuses from the same litter.

It should be noted that necropsy was performed on Day 21 post-coitum. At this time point, ossification of the skeleton should be almost complete. As such the higher incidence of unossified metatarsals and/or unossified sternebrae noted in all treated groups was regarded as treatment related. The exact cause of the delayed ossification is unknown. The (necrotizing) inflammation of the small intestine seen in the mothers may have resulted in impaired absorption of nutrition factors necessary for bone calcification. However, there were no data (like blood parameters) available from this study to substantiate this hypothesis.

No toxicologically significant changes were noted in any of the remaining developmental parameters investigated in this study (i.e. litter size, sex ratio,external, visceral and skeletal developmental malformations, external and visceral developmental variations). NOAEL maternal toxicity:

Based on the Granulocytic (necrotizing) inflammation in the ileum and granulomas with or without necrosis in the mesenteric lymph nodes in females starting at 3 mg/kg, no NOAEL for maternal toxicity could be derived. These findings are in agreement with the results obtained in other repeated dose studies for this substance.

NOAEL developmental toxicity:

The study report concluded that on the basis of the observed incidence of unossified metatarsal #1 also at the lowest dose of 3 mg/kg, no developmental NOAEL for Etherdiamine C13i/acetate could be derived. However, there are some additional comments that can be made to this:

- None of these increased incidences of delayed ossifications was statistically significant.

- The effects as such is not severe (classified as variation) and are also commonly seen in control groups. Higer incidences of such delayed ossifications are often seen at levels that result to maternal toxicity, and these delays are considered to be rapidly catching up after birth under normal feeding conditions.

- When a delayed development is noted (secondary to maternal toxicity), which is plausible considering the lower fetal body weights observed in the high dose fetuses, the evaluation for skeletal variation in principal shows a pattern of delayed ossifications. It seems that the data in this study point a narrow impact on the ossification of metatarsal #1 alone which is also visible at lower dose levels, but do not indicate a delayed ossification of other structures that would also be expected. For example, the reported proportion of unossified sternebrae was even lower in the 3 mg/kg group than in the control.

 

Looking at the total picture, there is a clear dose related effects on maternal health and nutritional status, already visible from the lowest dose, that clearly has a dose related effect on fetal development as shown by decreased fetal BW and delayed fetal growth. For both a dose effect relation is visible, and effects certainly clear at the highest dose level.

However, for the low (and mid-dose) levels the effects seem to show increased incidence of delayed ossification as observed compared to control specifically of the metatarsalis, however not so clear as for the high dose, and adversity might be debatable.

Based on the arguments above, we consider the reported small, statistically non-significant, increased incidence of unossified metatarsal #1 as observed at 3 mg/kg bw, as not adverse. The NOAEL for developmental toxicity therefore is set at 3 mg/kg bw.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
3 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
Available data are derived from recent, high quality and valid study showing consistent results.
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

In a prenatal developmental toxicity study rats were exposed to 0, 3, 7 or 15 mg/kg bw/d of the test substance once daily by oral gavage from days 6 to 20 post-coitum inclusive. The control group received the vehicle propylene glycol. Examinations were according to OECD 414 with additional histopathology of the stomach, small intestine and mesenteric lymph nodes.

 

Two high dose females at 15 mg/kg were euthanized moribund on Days 15 and 17 post-coitum, respectively. Toxicologically relevant clinical signs (lethargy, hunched posture, piloerection, and lean appearance) were noted at 15 mg/kg only. Treatment at 7 and 15 mg/kg resulted in reduced body weights, body weight gains, body weight corrected for uterine weight, and food consumption.

Treatment related adverse morphological alterations were present in the small intestine and the draining mesenteric lymph node of animals in all treated groups. At necropsy, a thickened wall of the duodenum, ileum and/or jejunum was noted which at the microscopic level correlated with (foamy) macrophages in the lamina propria and/or granulocytic (necrotizing) inflammation of the duodenum, jejunum and ileum. In general, these changes were dose related with the highest incidence and severity seen in the ileum. Enlarged mesenteric lymph nodes recorded at necropsy correlated with granulomas with or without necrosis (up to marked), extranodal inflammation with or without peritonitis, increased incidence, severity of increased macrophage foci, increased cellularity of cortex and/or medullary cords. Some of these findings were also noted in the 2 high dose females sacrificed moribund after 9 or 12 treatment days.

Based on the morphological data it is most likely that the (necrotizing) inflammation in the intestine caused by exposure to the test substance spreads to the draining mesenteric lymph nodes with possible breakthrough of granulomas into the abdominal cavity. Treatment at 15 mg/kg resulted in significantly lower fetal body weights (both sexes).This is most likely related to the reduced food consumption and body weight loss seen in the mothers.

 

A higher incidence of unossified metatarsal #1 was noted for all treated groups. The mean litter proportion was 8.6%, 10.2% and 18.9% in the 3, 7 and 15 mg/kg groups, respectively, as compared to 2.9% in the control group. None of these changes was statistically significant. It should, however, be noted that the litter proportion for the high dose group was in actuality even higher, since the 7 dead fetuses from litter 77 were not included in the group mean count.

There was also a trend towards a higher incidence of unossified sternebae in the 7 and 15 mg/kg groups. The mean litter proportion of unossified sternebra #5 was 1.6%, 4.5% and 4.6% in the 3, 7 and 15 mg/kg groups, respectively, as compared to 2.4% in the control group. In addition, unossified sternebra #2 with or without unossified vertebral centra and reduced ossification of the pubis was noted for 3 fetuses from the same litter.

 

It should be noted that necropsy was performed on Day 21 post-coitum. At this time point, ossification of the skeleton should be almost complete. As such the higher incidence of unossified metatarsals and/or unossified sternebrae noted in all treated groups was regarded as treatment related. The exact cause of the delayed ossification is unknown. The (necrotizing) inflammation of the small intestine seen in the mothers may have resulted in impaired absorption of nutrition factors necessary for bone calcification. However, there were no data (like blood parameters) available from this study to substantiate this hypothesis.

 

No toxicologically significant changes were noted in any of the remaining developmental parameters investigated in this study (i.e. litter size, sex ratio,external, visceral and skeletal developmental malformations, external and visceral developmental variations).

 

NOAEL maternal toxicity:

Based on the Granulocytic (necrotizing) inflammation in the ileum and granulomas with or without necrosis in the mesenteric lymph nodes in females starting at 3 mg/kg, no NOAEL for maternal toxicity could be derived. These findings are in agreement with the results obtained in other repeated dose studies for this substance.

NOAEL developmental toxicity:

The study report concluded that on the basis of the observed incidence of unossified metatarsal #1 also at the lowest dose of 3 mg/kg, no developmental NOAEL for Etherdiamine C13i/acetate could be derived. However, there are some additional comments that can be made to this:

- None of these increased incidences of delayed ossifications was statistically significant.

- The effects as such is not severe (classified as variation) and are also commonly seen in control groups. Higer incidences of such delayed ossifications are often seen at levels that result to maternal toxicity, and these delays are considered to be rapidly catching up after birth under normal feeding conditions.

- When a delayed development is noted (secondary to maternal toxicity), which is plausible considering the lower fetal body weights observed in the high dose fetuses, the evaluation for skeletal variation in principal shows a pattern of delayed ossifications. It seems that the data in this study point a narrow impact on the ossification of metatarsal #1 alone which is also visible at lower dose levels, but do not indicate a delayed ossification of other structures that would also be expected. For example, the reported proportion of unossified sternebrae was even lower in the 3 mg/kg group than in the control.

 

Looking at the total picture, there is a clear dose related effects on maternal health and nutritional status, already visible from the lowest dose, that clearly has a dose related effect on fetal development as shown by decreased fetal BW and delayed fetal growth. For both a dose effect relation is visible, and effects certainly clear at the highest dose level.

However, for the low (and mid-dose) levels the effects seem to show increased incidence of delayed ossification as observed compared to control specifically of the metatarsalis, but not as clear as for the high dose, and adversity might be debatable.

Based on the arguments above, we consider the reported, statistically non-significant increased incidences of unossified metatarsal #1 as observed at 3 mg/kg bw, as not adverse. The NOAEL for developmental toxicity therefore is set at 3 mg/kg bw.


Justification for selection of Effect on developmental toxicity: via oral route:
Study evaluating developmental toxicity of high validity.

Justification for selection of Effect on developmental toxicity: via inhalation route:
Likelihood of exposures via inhalation is low considering use and physical properties: The substance is very corrosive and its use is limited to industrial settings under controlled conditions. Also the high boiling point (> 300 °C) and low vapour pressure (0.005 Pa at 25°C) indicate that the potential for inhalation is not significant to justify this study.

Justification for selection of Effect on developmental toxicity: via dermal route:
Substance is corrosive. Effects will be characterized by local corrosive effects that are related to duration, quantity and concentration, rather than by systemic toxicity due to dermal uptake. Related to corrosive properties, exposures are likely to be limited in view of protective measures taken.

Justification for classification or non-classification

Study ofEtherdiamine C13i/acetate in a prenatal developmental toxicity study in rats has not shown reproductive or teratogenic effects. Developmental effects characterised as increased incidence of delayed ossification were only visible at maternal toxic dose levels, and are considered secondary to that.Further, in view of limited exposures, lack of effects on reproductive organs observed in repeated dose studies, and lack of effects on reproduction seen in reproduction and developmental toxicity studies on comparable fatty amine-like substances, there are no concerns for reproduction toxic effects by Etherdiamine C13i/acetate, and no classification for reproduction toxicity is required.