Registration Dossier

Administrative data

Key value for chemical safety assessment

Toxic effect type:
dose-dependent

Effects on fertility

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

Effects on developmental toxicity

Description of key information

The developmental toxicity of diethylhydroxylamine was evaluated in rats according to OECD Guideline 414. Diethylhydroxylamine was administered by oral gavage on gestation days 6 to 15. Maternal toxicity included decreased body weight and food consumption at 393 and 568 mg/kg/day. No evidence of developmental toxicity was observed at any dose level.

No teratogenic effects were detected in mice exposed to 8.9 ppm on days 6-17 of gestation. While negative, this study is compromised by technical and protocol deficiencies.

Link to relevant study records

Referenceopen allclose all

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
no
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
TEST ORGANISMS:
- Source: Charles River Laboratories, Inc., (Portage, Michigan)
- Age: approximately 71 days 
- Weight : from 216 g to 284 g on day 0 of gestation
- Fasting period before study: none
- Housing: were individually in clean, wire-mesh cages suspended above cage-board
- Diet (ad libitum): PMI Feeds, Inc.m Certified Rodent LabDiet© 5002
- Water (ad libitum): municipal water
- Acclimation period:

ENVIRONMENTAL CONDITIONS
- Temperature: 71.0°F to 72.4°F
- Humidity (%): 29.0 to 48.9
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
- Concentration in vehicle: 17.48, 78.7 and 113.6 mg/ml
- Total volume applied: 5 ml/kg
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The dosing formulations of the test article in deionized (DI) water were analyzed to determine the test article concentration by ultraviolet (UV) spectroscopy at 195 nm. The compound response was considered to be linear from 20.34 to 210.4 µg/mL (concentrations uncorrected for purity).
The dosing formulations were homogeneous with respect to the test article. The mean concentrations for all strata ranged from 98.2 to 102% of the target dose concentrations. The relative standard deviations (RSD) were 1.6, 0.75, and 0.65% for the three groups, indicating uniform dispersai of the test article in DI water.
The formulations were stored at room temperature for 8 days and then analyzed to assess stability. The formulations for Groups 3 and 4 were stable for at least 8 days. The Group 2 formulation was stable for at least 5 days; the 8-day concentration was slightly higher than the SOP specified upper limit.
The formulations that were analyzed and used for dose administration had concentrations within 10% of the target dose concentrations.
Details on mating procedure:
- Impregnation procedure: cohoused
- M/F ratio per cage: 1/1
- Length of cohabitation: until copulation
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility.
- Further matings after two unsuccessful attempts: no
- Verification of same strain and source of both sexes: yes
- Proof of pregnancy: vaginal plug and /or sperm in vaginal smear referred to as day 0 of pregnancy
- Any other deviations from standard protocol:
Duration of treatment / exposure:
from GD6 to GD15
Frequency of treatment:
Daily
Duration of test:
sacrifice on GD20
No. of animals per sex per dose:
25
Control animals:
yes, concurrent vehicle
Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: from day 0 through 20 of gestation (prior to test article administration during the dosing period). Animals were also observed for signs of toxicity approximately one hour following dosing.

BODY WEIGHT: Yes
- Time schedule for examinations: gestation days 0, 6-16 (daily) and 20

FOOD CONSUMPTION : Yes
- Time schedule for examinations: gestation days 0, 6-16 (daily) and 20

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 20
- Organs examined: uterus and ovaries

OTHER:
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other:
Fetal examinations:
- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: all per litter
- Skeletal examinations: Yes: all per litter
- Head examinations: Yes: all per litter
Statistics:
One-way ANOVA with Dunnett's test:
Corpora Lutea, Total Implantations, Fetal Body Weights, Maternal Body Weights and Weight Changes, Maternal Net Body Weight Changes and Gravid Uterine Weights, Food Consumption

Kruskal-Wallis test with Mann-Whitney U test:
Litter Proportions of Intrauterine Data (Considering the Litter, Rather than the Fetus, as the Experimental Unit)
Indices:
Postimplantation Loss/Litter
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
All animals survived to the scheduled necropsy on gestation day 20.
Treatment-related clinicat findings one hour following dosing in the 393 and 568
mg/kg/day groups included tan matting around the mouth and/or neck and
salivation between gestation days 9 and 15. Salivation was also noted at the time
of dosing in the 393 and 568 mg/kg/day groups generally beginning on gestation
day 12 and continuing through gestation day 15. In addition, single occurrences
of salivation were noted for two females in the 87.4 mg/kg/day group. However,
no other test article-related clinical findings were observed at this dose level, so
no relationship to treatment was evident. Haïr loss on various body surfaces was
noted in the 393 and 568 mg/kg/day groups generally from gestation day 7 through
study termination (gestation day 20). Other clinicat findings in all of the treated
groups occurred similarly in the control group, in single animals or in a manner
that was not suggestive of a relationship to treatment.
Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
ln the 568 mg/kg/day group, a statistically significant (p<0.01) mean body
weight loss was noted during gestation days 6-9. During this interval, 17 of 24
gravid females in this group lost between 1 and 18 grams of body weight. During
the remainder of the treatment period (gestation days 9-12 and 12-16), mean body
weight gains in the 568 mg/kg/day group were comparable to the control group
values. When the overall treatment period (gestation days 6-16) was evaluated,
mean body weight gain in this group was reduced relative to the control group
value; the difference was statistically significant (p < 0.01). During the
post-treatment period (gestation days 16-20), mean body weight gain in the 568
mg/kg/day group was similar to the control group value. Mean body weight from
gestation days 8 to 16, net body weight and net body weight gain in this group
-20-
WIL-160083
were slightly decreased compared to the control group values; the differences were
generally statistically significant (p < 0.01 or p < 0.05). Mean gravid uterine
weight in the 568 mg/kg/day group was comparable to the control group value.
A statistically significant (p < 0. 01) reduced mean body weight gain was noted
in the 393 mg/kg/day group during gestation days 6-9. During gestation days
9-12, mean body weight gain in this group was comparable to the control group
value. Du ring the remainder of the treatment period (gestation days 12-16), mean
body weight gain in the 393 mg/kg/day group was slightly reduced relative to the
control group value; the difference was statistically significant (p < 0.05).
However, a similar decrease in body weight gain during this interval was not
observed in the 568 mg/kg/day group; therefore, no relationship to treatment was
apparent. When the overall treatment period (gestation days 6-16) was evaluated,
mean body weight gain in this group was reduced when compared to the control
group value; the difference was statistically significant (p < 0.01). During the
post-treatment period (gestation days 16-20), mean body weight gain in the 393
mg/kg/day group was similar to the control group value. Mean body weights from
gestation days 7 to 16, net body weight and net body weight gain in this group
were slightly decreased relative to the control group values; the differences were
generally statistically significant (p < 0.01 or p < 0.05). Mean gravid uterine
weight in the 393 mg/kg/day group was comparable to the control group value.
Mean body weights, body weight gains, gravid uterine weight, net body
weight and net body weight gain in the 87.4 mg/kg/day group were unaffected by
test article administration. Values in this group were comparable to the control
group values; differences were slight and were not statistically significant.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Food consumption, evaluated as g/animal/day and g/kg/day, was reduced in
the 393 and 568 mg/kg/day groups during the entire treatment period (gestation
days 6-9, 9-12, 12-16 and 6-16). The differences from the control group were
statistically significant (p < 0. 01 or p < 0. 05). Du ring the post-treatment period,
food consumption in these groups was comparable to the control group.
Food consumption in the 87.4 mg/kg/day group was unaffected by treatment
with the test article. The only statistically significant (p < 0.05) difference from
the control group was a slight reduction (g/kg/day) in food consumption in this
group during the post-treatment period (gestation days 16-20). The g/animal/day
value was similar to the control group value, and no relationship to treatment was
apparent.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Description (incidence and severity):
At the scheduled necropsy on gestation day 20, 3 of the 25 females in the 393
mg/kg/day group and 12 of the 25 females in the 568 mg/kg/day group had hair
loss on multiple body surfaces (including the ventral abdominal, lateral abdominal,
ventral thoracic, hindlimbs and forelimbs). No test article-related internai findings
were noted at any dose level. In the control and 568 mg/kg/day groups, one
female each (nos. 66878 and 66918, respectively) had dilated renal pelves; female
no. 66918 also had a cystic kidney. Female no. 66888 in the 87.4 mg/kg/day
group had white areas and white precipitate in the kidneys. All other females were
intemally normal.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Total litter losses by resorption:
no effects observed
Early or late resorptions:
no effects observed
Changes in pregnancy duration:
not examined
Changes in number of pregnant:
no effects observed
Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
CLINICAL EXAMINATIONS DURING PREGNANCY
Treatment-related clinical findings noted in the 393 and 568 mg/kg/day  groups included hair loss on various body surfaces, tan matting around  the mouth and/or salivation. These findings were observed at the time of  dosing and/or one hour following dosing.

MATERNAL MORTALITY
All maternal animals survived to the scheduled necropsy on gestation day 20. 

MATERNAL BODY WEIGHT
During gestation days 6-9, a reduced mean body weight gain occurred in the 393 mg/kg/day group and a mean body weight loss occurred in the 568 mg/kg/day group; the differences from the control group were statistically significant. During gestation days 9-12, mean body weight gains in the 393 and 568 mg/kg/day groups were comparable to the control group values. During the remainder of the treatment period (gestation days 12-16), mean body weight gains were reduced (statistically significant) in the 393 mg/kg/day group and were similar to the control group in the 568 mg/kg/day group. However, the decrease in body weight gain in the 393 mg/kg/day group during this interval was not considered to be related to treatment because a corresponding decrease was not noted in  the 568 mg/kg/day group. Mean body weight gains in the 393 and 568  mg/kg/day groups were comparable to the control values during the  post-treatment period (gestation days 16-20). Mean body weights in these  groups were reduced (statistically significant) beginning on gestation  day 7 (393 mg/kg/day) or 8 (568 mg/kg/day) and continuing through  gestation day 16. Mean net body weights and net body weight gains in  these groups were slightly decreased relative to the control group  values. 

FOOD CONSUMPTION
Food consumption, evaluated as g/animal/day and g/kg/day, was reduced in the 393 and 568 mg/kg/day groups throughout the entire treatment period (gestation days 6-9, 9-12 and 12-16); the differences from the control group were statistically significant. During the post-treatment period (gestation days 16-20), food consumption in these groups was comparable  to the control group. Food consumption in the 87.4 mg/kg/day group was  unaffected by treatment with the test article.

ORGAN WEIGHT
Mean gravid uterine weights in the 393 and 568 mg/kg/day groups were  similar to the control group value. Body weight data in the 87.4  mg/kg/day group was unaffected by test article administration.

MACROSCOPIC EXAMINATION
At the scheduled necropsy on gestation day 20, no test article-related internal findings were observed at any dose level.
Dose descriptor:
NOAEL
Effect level:
393 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
body weight and weight gain
Abnormalities:
no effects observed
Fetal body weight changes:
no effects observed
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
no effects observed
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Extemal malformations were observed in one fetus each in the control, 393
and 568 mg/kg/day groups. Control group fetus no. 66833-06 had vertebral
agenesis (at the skeletal examination, ail vertebrae posterior to lumbar no. 5 were
absent). Fetus no. 66854-03 in the 393 mg/kg/day group had maxillary
micrognathia. ln the 568 mg/kg/day group, fetus no. 66901-12 had
microphthalmia, anophthalmia, micromelia, adactyly and thoracogastroschisis. No
other extemal malformations were noted.
No extemal developmental variations were observed in fetuses at any dose
level.
Skeletal malformations:
no effects observed
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Soft tissue malformations were observed in 1(1), 1(1), 2(2) and 1(1) fetuses
(litters) in the control, 87.4, 393 and 568 mg/kg/da y groups, respectively. One
fetus each in the control, 87.4 and 568 mg/kg/day groups (nos. 66857-11,
66909-14 and 66901-12, respectively) had a heart anomaly with an associated great
vessel anomaly. However, since the heart and great vessel anomalies in each of
the affected fetuses was dissimilar and were noted similarly in the control group,
no relationship to treatment was evident. ln addition, fetus no. 66857-11 in the
control group had situs inversus and pulmonary lobular dysgenesis and fetus no.
66901-12 in the 568 mg/kg/day group had an absent diaphragm and a missing
adrenal gland. In the 393 mg/kg/day group, two females (nos. 66854-03 and
66917-01) each had situs inversus. No other soft tissue malformations were noted.
The only soft tissue variation observed was distended ureters in fetus no.
66872-07 in the 87.4 mg/kg/day group.
One fetus (no. 66927-15) in the 393 mg/kg/day group had reddened testes.
This finding was not classified as either a malformation or variation and was not
included in any tabulation.
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
EXAMINATION OF FETUSES AND UTERINE CONTENT
Intrauterine growth and survival were unaffected by test article administration at any dose level.  Parameters evaluated included postimplantation loss, live litter size,  mean fetal body weights, fetal sex ratios and the mean numbers of corpora  lutea and implantation sites. Fetuses (litters) available for morphological evaluation numbered 354(24), 326(22), 353(23) and 356(24) in the control, 87.4, 393 and 568 mg/kg/day groups, respectively. Malformations were observed in 2(2), 1(1), 2(2) and 3(3) fetuses (litters) in these same respective dose groups and were considered to be spontaneous in origin. No developmental variants were noted in fetuses in the treated groups that were considered to be related to treatment with the test article.
Dose descriptor:
NOAEL
Effect level:
568 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Abnormalities:
effects observed, non-treatment-related
Developmental effects observed:
no
Conclusions:
In conclusion, maternal toxicity was expressed by inhibition of body weight gain and food consumption at dose levels of 393 and 568 mg/kg/day. No maternal toxicity was evident at a dose level of 87.4 mg/kg/day. No developmental toxicity was apparent at any dose level. Based on the results of this study, the NOAEL (no observable adverse effect level) for maternal toxicity was considered to be 87.4 mg/kg/day, and the NOAEL for developmental toxicity was considered to be 568 mg/kg/day.
Executive summary:

The potential maternal toxicity and developmental toxicity of diethylhydroxylamine were evaluated. Diethylhydroxylamine in the vehicle, deionized water, was administered to three groups of 25 bred Crl:CD®(SD)BR rats once daily from gestation days 6 through 15. Dosage levels were 87.4, 393 and 568 mg/kg/day administered at a dose volume of 5 ml/kg. A concurrent control group composed of 25 bred females received the vehicle, deionized water, on a comparable regimen at 5 ml/kg. The route of administration was oral by gastric intubation. Clinical observations, body weights and food consumption were recorded. On gestation day 20, a laparohysterectomy was performed on ail animals. The uteri and ovaries were examined and the numbers of fetuses, early and late resorptions, total implantations and corpora lutea were recorded. Mean gravid uterine weights and net body weight changes were calculated for each group. The fetuses were weighed, sexed and examined for external, soft tissue and skeletal malformations and variations. All maternal animals survived to the scheduled necropsy on gestation day 20. Treatment-related clinical observations noted at the time of dosing and/or one hourfollowing dosing in the 393 and 568 mg/kg/day groups included tan matting around themouth and/or neck, salivation and hair loss on various body surfaces. During the first three days of dosing, a reduced mean body weight gain occurred in the 393 mg/kg/day group and a mean body weight loss occurred in the 568 mg/kg/day group. Food consumption was reduced in the 393 and 568 mg/kg/day groups throughout the treatment period. Body weight data and food consumption in the 87.4 mg/kg/day group were unaffected by treatment with the test article. No test article-related internal findings were observed at any dose level. Intrauterine growth and survival were unaffected by test article administration at any dose level. The fetal malformations and developmental variations observed in the treated groups were considered to be spontaneous in origin. Based on the results of this study, the NOAEL (no observable adverse effect level) for maternal toxicity was considered to be 87.4 mg/kg/day and the NOAEL for developmental toxicity was considered to be 568 mg/kg/day.


Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study well documented, meets generally accepted scientific principles, acceptable for assessment
Principles of method if other than guideline:
Developmental toxicity range-finding study
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
TEST ORGANISMS:
- Source: Charles River Laboratories, Inc., (Portage, Michigan)
- Age: approximately 71 days
- Weight : from 216 g to 284 g on day 0 of gestation
- Fasting period before study: none
- Housing: were individually in clean, wire-mesh cages suspended above cage-board
- Diet (ad libitum): PMI Feeds, Inc.m Certified Rodent LabDiet© 5002
- Water (ad libitum): municipal water
- Acclimation period:

ENVIRONMENTAL CONDITIONS
- Temperature: 67.4°F to 73.0°F
- Humidity (%): 37.2 to 49.4
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
other: deionized water
Details on exposure:
- Total volume applied: 5 ml/kg
Analytical verification of doses or concentrations:
no
Details on mating procedure:
Females were placed in a suspended wire-mesh cage with a resident  untreated male. Positive evidence of mating was confirmed by the presence of a copulatory  plug or the presence of sperm in vaginal smear, checked daily. The day on  which evidence of mating  was identified was termed day 0 of gestation.
Duration of treatment / exposure:
from GD6 to GD15
Frequency of treatment:
daily
Duration of test:
sacrifice on GD20

No. of animals per sex per dose:
8
Control animals:
yes, concurrent vehicle
Details on study design:
Sex: female
Duration of test: sacrifice on GD20
Maternal examinations:
- Maternal body weight: recorded on days 0, 6 to 16 (daily) and 20.
- Food consumption: recorded on days 0, 6 to 16 (daily) and 20.
- Clinical observations: twice daily from day 0 through day 20
- Maternal mortality: twice daily from day 0 through day 20
- Organ weights: gravid uteri
- Histopathology: Thoracic, abdominal and pelvic cavities were opened by  a ventral midline incision and the contents examined.
Ovaries and uterine content:
- Examination of uterine content: laparohysterectomy on day 20, early and  late resorption, total implantations, number of corpora lutea
Fetal examinations:
- Examination of fetuses: number of fetuses, sex ratio, external  malformations
Statistics:
- One-way ANOVA with Dunnett's test applied to corpora lutea, total  implantations, viable fetuses, fetal body weights, maternal body weights,  gravid uterine weights and food consumption.
- Kruskall-Wallis test with Mann-Whitney U test for applied to litter  proportions of intrauterine data (considering the litter rather than the  fetus).
Clinical signs:
effects observed, treatment-related
Dermal irritation (if dermal study):
not examined
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Immunological findings:
not examined
Histopathological findings: neoplastic:
not examined
Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
Five females in the 1000 mg/kg/day group died between gestation days 10 and 14. The remaining three females in this group were euthanized and discarded (without necropsy) on gestation day 16 or 17, at the request of the sponsor. All other animals survived to the scheduled necropsy on gestation day 20 No test article-related internal findings were noted at any dose level.  Clinical observations related to treatment with the test article included  salivation, animals rubbing their muzzles on the cage floor, tan  matting/material around the mouth and neck and drooping eyelids at dose  levels of 500, 750 and/or 1000 mg/kg/day at the time of dosing or one  hour following dosing.  Mean body weight losses and reduced food consumption were observed in the  500, 750 and 1000 mg/kg/day groups during gestation days 6-9.  Food consumption remained reduced in the 750 mg/kg/day group during gestation days 9-12 and 12-16.
Dose descriptor:
NOAEL
Effect level:
250 mg/kg bw/day
Basis for effect level:
clinical signs
body weight and weight gain
Fetal body weight changes:
effects observed, treatment-related
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
no effects observed
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
A slighltly reduced mean fetal body weight was observed in the 750  mg/kg/day group. Intrauterine growth and survival were unaffected by test article  administration at any other dose level evaluated. The only external  malformations observed were anuria in in one control group fetus and  microphtalmia and craniorachischisis in one 500mg/kg/day group fetus. No  external developmental variations were observed in fetuses at any dose  level.
Abnormalities:
not specified
Developmental effects observed:
not specified
Conclusions:
Maternal toxicity was expressed by mortalities at a dose level of 1000 mg/kg/day and changes in the clinical condition of animals and inhibition of body weight gain and food consumption at dose levels of 500, 750 and 1000 mg/kg/day. No maternal toxicity was observed at a dose level of 100 and 250 mg/kg/day. No treatment related developmental toxicity was observed at dose levels of 100, 250, 500 and 750 mg/kg/day.
Executive summary:

The potential maternal toxicity and developmental toxicity of diethylhydroxylamine were evaluated. Diethylhydroxylamine in the vehicle, deionized water, was administered to five groups of eight bred Crl: CD(SD)BR rats once daily from gestation days 6 through 15. Dosage levels were 100, 250, 500, 750 and 1000 mg/kg/day administered at a dose volume of 5 ml/kg. A concurrent control group composed of eight bred females received the vehicle, deionized water, on a comparable regimen at 5 ml/kg. The route of administration was oral by
gastric intubation. Clinical observations, body weights and food
 consumption were recorded. On gestation day 20, a laparohysterectomy was performed on all surviving animals (Groups 1-5). The uteri and ovaries were examined and the numbers of fetuses, early and late resorptions, total implantations and corpora lutea were recorded. Mean gravid uterine weights and net body weight changes were calculated for each group. The fetuses were weighed, sexed and examined for external malformations and variations.
Five females in the 1000 mg/kg/day group died between gestation days 10
 and 14. The remaining three females in this group were euthanized and discarded (without necropsy) on gestation day 16 or 17, at the request of the sponsor. All other animals survived to the scheduled necropsy on gestation day 20; no test article-related internal findings were noted at any dose level. Clinical observations related to treatment with the test article included salivation, animals rubbing their muzzles on the cage floor, tan matting/material around the mouth and neck and drooping eyelids at dose levels of 500, 750 and/or 1000 mg/kg/day at
the time of dosing or one hour following dosing. Mean body weight losses
 and reduced food consumption were observed in the 500, 750 and 1000 mg/kg/day groups during gestation days 6-9. Food consumption remained reduced in the 750 mg/kg/day group during gestation days 9-12 and 12-16.
A slightly reduced mean fetal body weight was observed in the 750
 mg/kg/day group. Intrauterine growth and survival were unaffected by test article administration at any dose level evaluated.
The only external malformations observed were anuria in one control
 group fetus and microphtalmia and craniorachischisis in one 500mg/kg/day group fetus. No external developmental variations were observed in fetuses at any dose level.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
568 mg/kg bw/day
Study duration:
subacute
Experimental exposure time per week (hours/week):
7
Species:
rat
Quality of whole database:
GLP guideline study
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

Rat study

Developmental or Reproductive Toxicity/ Female CD (SD) female rats were dosed once daily from GD6-GD15 with 87.4, 393 or 568 mg/kg diethylhydroxylamine by gavage (Stump, 1997a). Animals were sacrificed on GD20. During gestation days 6 -9, a reduced mean body weight gain occurred in the 393 mg/kg/day group and a mean body weight loss occurred in the 568 mg/kg/day group; the differences from the control group were statistically significant. Food consumption, evaluated as g/animal/day and g/kg/day, was reduced in the 393 and 568 mg/kg/day groups throughout the entire treatment period (gestation days 6-9, 9-12 and 12-16); the differences from the control group were statistically significant. All maternal animals survived to the scheduled necropsy on gestation day 20. Intrauterine growth and survival were unaffected by test article administration at any dose level. Parameters evaluated included postimplantation loss, live litter size, mean fetal body weights, fetal sex ratios and the mean numbers of corpora lutea and implantation sites. Fetuses (litters) available for morphological evaluation numbered 354(24), 326(22), 353(23) and 356(24) in the control, 87.4, 393 and 568 mg/kg/day groups, respectively. In conclusion, maternal toxicity was expressed by inhibition of body weight gain and food consumption at dose levels of 393 and 568 mg/kg/day. No maternal toxicity was evident at a dose level of 87.4 mg/kg/day. No developmental toxicity was apparent at any dose level. Based on the results of this study, the NOAEL (no observable adverse effect level) for maternal toxicity was considered to be 87.4 mg/kg/day, and the NOAEL for developmental toxicity was considered to be 568 mg/kg/day.

In a range-finding developmental toxicity study (Stump, 1997b), diethylhydroxylamine was administered by gastric intubation at dose levels of 0, 100, 250, 500, 750 and 1000 mg/kg/d to 5 groups of 8 bred CD rats once daily from GD 6 through 15. On GD 20, a laparohysterectomy was performed on all surviving animals. Five females in the 1000 mg/kg/d group died between GD10 and GD 14, and the remaining 3 females were euthanized on GD 16 or 17. Clinical observation included salivation, animals rubbing their muzzles on the cage floor, tan matting/material around the mouth and neck and drooping eye lids at dose levels of 500, 750 and/or 1000 mg/kg/d. Mean body weight losses and reduced food consumption were observed in the 500, 750, and 1000 mg/kg/d group during GD 9-12 and 12-16. A slightly reduced mean fetal body weight was observed in the 750 mg/kg/d group. Intrauterine growth and survival were unaffected by diethylhydroxylamine administration. The only external malformations observed were anury in one control group fetus and microphthalmia and craniorachischisis in one 500 mg/kg/d group fetus. No external developmental variations were observed.

Mouse study

Pregnant female mice were exposed from Days 6 to 17 of pregnancy to 8.9 +/- 2.0 ppm of diethylhydroxylamine and 14.3 +/- 2.0 ppm of nitroethane (Beliles et al., 1978). Exposures were for 8.25 +/- 2.25 hours/day including weekends. The mice were also exposed continuously for 24 hours a day to the vapor of diethylamine hydrogen sulfite. The exposure produced no effect on the dams compared to the control group. There was no evidence of compound-induced terata, variation in sex ratio, embryotoxicity, or inhibition of fetal growth and development. However, technical and protocol deficiences in this study preclude the use of this data to unequivocally evaluate the developmental toxicity potential of DEHA.

Toxicity to reproduction: other studies

Link to relevant study records
Reference
Endpoint:
toxicity to reproduction: other studies
Type of information:
experimental study
Adequacy of study:
key study
Study period:
09 March 2018 - 06 August 2018
Reliability:
1 (reliable without restriction)
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
other: OECD 408
GLP compliance:
yes (incl. certificate)
Type of method:
in vivo
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: breeder: Charles River Laboratories Italia, Calco, Italy
- Age: on the first day of treatment, the animals were 5 to 6 weeks old
- Mean body weight: on the first day of treatment, the males had a mean body weight of 199 g (range: 169 g to 229 g) and the females had a mean body weight of 157 g (range: 137 g to 180 g)
- Fasting period before study: no
- Housing: the animals were housed in twos or threes from the same sex and group, in polycarbonate cages with stainless steel lids (Tecniplast 2000P, 2065 cm²) containing autoclaved sawdust
- Diet: SSNIFF R/M-H pelleted diet (free access)
- Water: tap water filtered with a 0.22 µm filter (free access)
- Acclimation period: for a period of 11 days before the beginning of the treatment period.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2°C
- Humidity (%): 50 ± 20%
- Air changes (per hr): approximately 8 to 15 cycles/hour of filtered, non-recycled air
- Photoperiod (hrs dark / hrs light): 12 h/12 h

IN-LIFE DATES: 26 March 2018 to 06 August 2018.
Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
PREPARATION OF DOSING FORMULATIONS:
- Solution in the vehicle
- Concentration in vehicle: 10, 30 and 100 mg/mL
- Amount of vehicle (if gavage): 5 mL/kg/day.
Details on analytical verification of doses or concentrations:
Type of method: Gas Chromatography with FID detection (GC-FID)
Test item concentrations:
Once in Weeks 1, 4, 8 and 12, all within the acceptance criterion.
A sample was taken from control and test item dose formulations and analyzed using the validated method
Acceptance criterion:
Measured concentration = nominal concentration ± 10%

Homogeneity: not assessed, formulation was a solution.
Checked parameters, acceptance criteria and obtained results are detailed in the validation report

Stability: 5 days at room temperature.
Duration of treatment / exposure:
13 weeks followed by a 6-week treatment-free period
Frequency of treatment:
DAILY
Duration of test:
13 weeks of dosing.
Dose / conc.:
0 mg/kg bw/day (nominal)
Dose / conc.:
50 mg/kg bw/day (nominal)
Dose / conc.:
150 mg/kg bw/day (nominal)
Dose / conc.:
500 mg/kg bw/day (nominal)
No. of animals per sex per dose:
10 males and 10 females per group (+ 5 males and 5 females for treatment-free period in control and high dose groups).
Control animals:
yes, concurrent vehicle
Details on study design:
- Rationale for dose selection:
The dose levels were selected in agreement with the Sponsor, based on the results of previous toxicity studies: a 2-week dose-range finding study and a 28-day toxicity study (OECD 407), both performed in the same species.
In the 2-week dose-range finding study, three groups of Sprague-Dawley rats were treated with the test item at 100 mg/kg, 300 mg/kg and 1000 mg/kg by the oral route. At 1000 mg/kg/day, lower body weight gain was noted in males over the study period. Higher liver (+14% to +23%) and kidney (+10 to +22%) weights were recorded in both sexes when compared to controls. At 300 mg/kg/day, higher liver (+15%) and kidney (+10%) weights were noted in females only. At 100 mg/kg/day, no test item related changes were noted.

In the 28-day toxicity study, three groups of Sprague-Dawley rats received the test item at 20, 100 or 500 mg/kg/day by the oral route. No changes in body and organ weights were noted. At microscopic examination, the changes observed were considered to be incidental.

- Rationale for animal assignment: computerized randomization procedure.
Dose descriptor:
NOAEL
Effect level:
500 mg/kg bw/day (nominal)
Sex:
male/female
Basis for effect level:
other: Absence of adverse effect
Remarks on result:
not determinable due to adverse toxic effects at highest dose / concentration tested
Executive summary:

In the OECD 408 study (Modeste, 2018), the estrous cycle was determined over 21 or 14 consecutive days for all females at the end of the treatment or treatment-free period, respectively. At the end of the treatment period, seminological investigations (count and motility) were performed on all males, and sperm morphology was determined in control and high-dose males. On completion of the treatment, epididymides, ovaries (including oviducts), testes and uterus (horns and cervix) were weighed. A microscopic examination was performed on epididymides, ovaries, prostate , seminal vesicles (including coagulation gland), testes, uterus and vagina from control and high-dose animals euthanized at the end of the treatment period. There were no effects on reproductive organ weight or at pathology investigation.

At sperm analysis, motility was unaffected but there were dose-related slightly lower percentage of morphologically normal epididymal sperm, lower number of epididymal and testicular sperm as well as lower daily sperm production rate at 150 and 500 mg/kg/day; dose-related higher mean percentages of sperm with abnormal or no flagellum was noted at all three dose levels. The magnitude of the changes being low and in absence of test item-related histopathological findings in the right testis and epididymis at the high-dose, none of these findings were considered to be adverse. They were fully reversible at the end of the 6-week treatment-free period, except for epididymal sperm count which still remained slightly lower than in controls.

The NOAEL of 500 mg/kg/j was set at the highest dose tested.

Justification for classification or non-classification

On the basis of the findings in the developmental toxicity studies in rat, DEHA does not warrant classification for reproductive and developmental toxicity according to Regulation (EC) No 1272-2008.