2-dibutylaminoethanol

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

05Mar2019 to 27Nov2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2020

Report date:

2020

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Version / remarks:

25 Jun 2018

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

2-dibutylaminoethanol

EC Number:

203-057-1

EC Name:

2-dibutylaminoethanol

Cas Number:

102-81-8

Molecular formula:

C10H23NO

IUPAC Name:

2-(dibutylamino)ethan-1-ol

Test material form:

other: liquid

Details on test material:

- Name of test material (as cited in study report): N,N-Dibutylethanolamine
- Physical state: liquid

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch number of test material: A028-2018
- Expiration date of the lot/batch: 09 Apr 2020
- Purity: 99.8 corr. area-%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room temperature
- The stability of the test substance under storage
conditions over the test period was guaranteed by the sponsor, and the sponsor holds this
responsibility.
- Solubility and stability of the test substance in the vehicle The stability of Dibutylethanolamine
in corn oil at room temperature for a period of 7 days was proven before the start of the
administration period

FORM AS APPLIED IN THE TEST
- The test substance was applied as an oily preparation

OTHER SPECIFICS
- Physical state/ appearance: Liquid/ yellowish, clear
- Homogeneity: Given

Test animals

Species:

rat

Strain:

Wistar

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH
- Age at study initiation: 10-12 weeks
- Weight at study initiation: 163.5 - 218.2
- Housing: During the study period, the rats were housed individually in Polycarbonate cages type III
supplied by TECNIPLAST, Hohenpeißenberg, Germany and Becker & Co., Castrop-Rauxel,
Germany (floor area about 800 cm²). Individual housing is preferred over group housing as the
close individual monitoring of food and water consumption as well as of clinical signs of toxicity
in pregnant animals is crucial during this period of increased sensitivity. In addition, the control
for signs of abortion or fetal loss can only be done in a reliable fashion with single-housed
animals
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: GD 0 - GD 6

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24°C
- Humidity (%): 45-65%
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

-IN-LIFE DATES; From: To:
Study initiation date: 04 Mar 2019
Experimental starting date:(arrival of 1st cohort of test animals) 05 Mar 2019
Supply of animals/ Beginning of acclimatization (GD 0) Beginning oftreatment/
End of acclimatization (GD 6)
End of treatment (GD 19)
Blood sampling / Sacrifice of the animals (GD 20)
1st cohort 05 Mar 2019 11 Mar 2019 24 Mar 2019 25 Mar 2019
2nd cohort 06 Mar 2019 12 Mar 2019 25 Mar 2019 26 Mar 2019
3rd cohort 07 Mar 2019 13 Mar 2019 26 Mar 2019 27 Mar 2019
Experimental completion date: (draft to QAU*) 27 Nov 2019
* QAU = Quality Assurance Unit

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
The test substance preparations were prepared at the beginning of the administration period
and thereafter at intervals, which took into account the period of established stability. The
preparations were kept at room temperature.
For the test substance preparations, the specific amount of test substance was weighed,
topped up with corn oil in a graduated flask and intensely mixed with a magnetic stirrer until it
was completely dissolved.

TEST GROUPS AND DOSES
Test group Dose [mg/kg bw/d] Concentration [g/100 mL] Volume [mL/kg bw/d] .
0: 0 [mg/kg bw/d] 0 [g/100 mL] 4a)
1: 20 [mg/kg bw/d] 0.50[g/100 mL] 4b)
2: 60 [mg/kg bw/d] 1.50 [g/100 mL] 4b)
3: 200[mg/kg bw/d] 5.00 [g/100 mL] 4b)
a) corn oil
b) Test substance preparations in corn oil

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Analytical verifications of the stability of the test substance in corn oil over a period of 7 days
at room temperature had been verified prior to the start of the study. Samples of the test
substance preparations were sent twice (at the beginning of administration and after the end
of study to the analytical laboratory for verification of the concentrations.

Details on mating procedure:

Impregnation procedure: purchased timed pregnant

The animals were paired by the breeder (“time-mated”); the day of evidence of mating
(= detection of vaginal plug/sperm) was referred to as GD 0. The animals arrived on the same
day (GD 0) at the experimental laboratory. The following day was designated as “GD 1”. The
animals were acclimated to the laboratory conditions between start of the study (beginning of
the experimental phase) and first administration (GD 6).

Duration of treatment / exposure:

The test substance preparations were administered to the animals once a day orally by gavage,
from implantation to one day prior to the expected day of parturition (GD 6 to GD 19), always
at approximately the same time in the morning. The animals of the control group were treated
with the vehicle (corn oil) in the same way. The volume administered each day was 4 mL/kg
body weight. The calculation of the administration volume was based on the most recent
individual body weight.

Frequency of treatment:

The test substance preparations were administered to the animals once a day orally by gavage,
from implantation to one day prior to the expected day of parturition (GD 6 to GD 19), always
at approximately the same time in the morning.

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (actual dose received)

Remarks:

control

Doses / concentrations 2

Dose / conc.:

20 mg/kg bw/day (actual dose received)

Remarks:

low dose

Doses / concentrations 3

Dose / conc.:

60 mg/kg bw/day (actual dose received)

Remarks:

mid dose

Doses / concentrations 4

Dose / conc.:

200 mg/kg bw/day (actual dose received)

Remarks:

high dose

No. of animals per sex per dose:

25

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale:
In a range-finding study (BASF project No. 10C0286/05S039), Dibutylethanolamine was
administered by gavage to groups of five non-pregnant female Wistar rats for 28-days at dose
levels of 0, 100 and 300 mg/kg body weight/day (mg/kg bw/d). Corn oil served as vehicle. A
dose of 300 mg/kg bw/d induced neurobehavioral changes in females from study day 10
onwards. Clinical findings consisted of tremors in all females, twitching in three and lateral
position in two out of five females. Furthermore, mortalities were seen in each one female on
study days 14 and 17. From study day 18 onwards, this dose level was reduced to 200 mg/kg
bw/d. In females, clonic convulsions occurred still in one animal on study day 18. Twitching
was observed in up to two females on study days 19 and 20. All mentioned findings occurred
between 15 min and 2 hours after administration but not thereafter. All other parameters were
not affected. At the lowest dose of 100 mg/kg bw/d, no relevant findings were observed.

In a rat maternal toxicity study (BASF project No. 10R0286/05R038), Dibutylethanolamine
was administered orally by repeated gavage to up to seven pregnant Wistar rats per test group
from gestation day (GD) 6 through GD 19. Doses of 200 and 70 mg/kg bw/d were selected
based on the findings observed in the above-mentioned range-finding study. At 200 mg/kg
bw/d, signs of systemic toxicity were observed. They consisted of a statistically significant
reduction of water (GD 6-8: 25% below control) and food (GD 6-13: 31%; GD 6-19: 12% below
control) consumption, a statistically significant decrease in body weight change (including a
body weight loss during GD 6-8: -1.4 g vs. 6.9 g in control) and a decrease in net weight change
(27% below control). At 70 mg/kg bw/d, no relevant, adverse findings were observed.
Based on the findings presented in the two studies above, 200 mg/kg bw/d was selected as
high-dose in the following prenatal development toxicity study to show some maternal toxicity
as stated in the respective OECD test guideline No. 414.
The following dose levels were chosen for the present prenatal
developmental toxicity study in Wistar rats:
20 mg/kg body weight/day: as low-dose level
60 mg/kg body weight/day: as mid-dose level
200 mg/kg body weight/day: as high-dose level #
The oral route was selected since this has proven to be suitable for the detection of a toxicological hazard.

# Because of severe clinical findings, such as twitching, convulsions, tremor and/or abdominal
position after treatment, in almost all animals, the high-dose group (200 mg/kg bw/d) was
sacrificed prematurely on GD 13/12/11 of the study. The animals were discarded without further
examinations. No organs or tissues were fixed, but the pregnancy status was evaluated.

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily
- During treatment period (GD 6-19) all rats were checked daily for any signs
of morbidity, pertinent behavioral changes and/or signs of overt toxicity before administration
as well as within 2 hours and within 5 hours after administration.
- A check was made twice a day on working days or once a day on Saturdays, Sundays or on
public holidays (GD 0-20).

BODY WEIGHT: Yes
All animals were weighed on GD 0, 1, 3, 6, 8, 10, 13, 15, 17, 19 and 20. The body weight
change of the animals was calculated based on the obtained results.
Corrected (net) body weight gain
Furthermore, the corrected body weight gain was calculated after terminal sacrifice (terminal
body weight on GD 20 minus weight of the unopened uterus minus body weight on GD 6).

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
The consumption of food was recorded for the intervals GD 0-1, 1-3, 3-6, 6-8, 8-10, 10-13,
13-15, 15-17, 17-19 and 19-20.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
-The consumption of water was recorded for the intervals GD 0-1, 1-3, 3-6, 6-8, 8-10, 10-13,
13-15, 15-17, 17-19 and 19-20.


POST-MORTEM EXAMINATIONS: Yes / No / No data
- Sacrifice on gestation day 20
- Organs examined: Necropsy

OTHER:
After the dams had been sacrificed, they were necropsied and assessed for gross pathology.
Thyroid glands (including parathyroid glands) were sampled.

Thyroid hormones
The concentrations of TSH were determined:
Total triiodothyronine (T3), Total thyroxine (T4), Thyroid stimulating hormone (TSH)
The following weights were determined in all dams sacrificed on schedule:
Thyroid glands (with parathyroid glands) (fixed)
Thyroid glands were weighed together (left and right).

Ovaries and uterine content:

After the dams had been sacrificed, they were necropsied and assessed for gross pathology.
Thyroid glands (including parathyroid glands) were sampled.
The uteri and the ovaries were removed and the following data were recorded:
Weight of the unopened uterus, Number of corpora lutea,
Number and distribution of implantation sites classified as: Live fetuses,
Dead implantations:
Early resorptions (only decidual or placental tissues visible or according to SALEWSKI
from uteri from apparently non-pregnant animals and the empty uterus horn in the
case of single horn pregnancy)
Late resorptions (embryonic or fetal tissue in addition to placental tissue visible)
Dead fetuses (hypoxemic fetuses which did not breathe spontaneously after the
uterus had been opened)
After the weight of the uterus had been determined, all subsequent evaluations of the dams
and the gestational parameters (except of gross pathology including organ sampling and
weights) were conducted.

Fetal examinations:

At necropsy each fetus was weighed, sexed, and external tissues and all orifices were
examined macroscopically. The sex was determined by observing the distance between the
anus and the base of the genitalia.
Furthermore, the viability of the fetuses and the condition of placentas, umbilical cords, fetal
membranes, and fluids were examined. The placentas were weighed and their individual
weights were recorded.
Thereafter, the fetuses were sacrificed by a subcutaneous injection of pentobarbital
(Narcoren®; dose: 0.1 mL/fetus).
The anogenital distance (defined as the distance from the center of the anal opening to the
base of the genital tubercle) measurements were conducted, using a measuring ocular, on all
liveborn fetuses.
After these examinations, approximately one half of the fetuses per dam were eviscerated,
skinned and fixed in ethanol; the other half was placed in Harrison’s fluid for fixation.

Soft tissue examination of the fetuses
The fetuses fixed in Harrison’s fluid were examined for any visceral findings according to the
method of BARROW and TAYLOR. After this examination these fetuses were discarded.

Skeletal examination of the fetuses
The skeletons of the fetuses fixed in ethanol were stained according to a modified method of
KIMMEL and TRAMMELL. Thereafter, the skeletons of these fetuses were examined under a
stereomicroscope. After this examination the stained fetal skeletons were retained individually.

Statistics:

Simultaneous comparison of all dose groups with the control group using theDUNNETT-test
(two-sided) for the hypothesis of equal means.:
Water consumptiona), food consumptiona), body weight, body weight change, corrected body
weight gain (net maternal body weight change), carcass weight, weight of unopened uterus,
number of corpora lutea, number of implantations, number of resorptions, number of live fetuses,
proportions of preimplantation loss, proportions of postimplantation loss, proportions of resorptions,
proportion of live fetuses in each litter, litter mean fetal body weight, litter mean placental weight,
anogenital distance, anogenital index
Pairwise comparison of eachdose group with the control group using FISHER'S EXACT test
(one-sided) for the hypothesis of equal proportions: Female mortality, females pregnantat terminal
sacrifice, number of litters with fetal findings.
Pairwise comparison of each dose group with the control group using the WILCOXONtest
(one-sided) for the hypothesis of equal medians.

Indices:

Conception rate, pre-impantation loss, post-implantation loss

Historical control data:

yes

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

For almost all females of test group 3 (200 mg/kg bw/d) severe clinical findings were recorded
during the treatment period (within the 2-hour examination interval after treatment):
twitching after treatment in 23 females from GD 7 onwards,
convulsion after treatment in 6 females from GD 10 onwards,
tremor after treatment in 3 females on GD 9 and 11,
abdominal position after treatment in 17 females from GD 8 onwards.
As mentioned above, all females of test group 3 were sacrificed prematurely on GD 13/12/11.
Furthermore, most females (23 out of 25) of the high-dose group and 6 females of the middose
group showed transient salivation during the treatment period. Salivation occurred in the
respective animals within the 2-hour examination interval after treatment (i.e. 0-2h) and was
initially observed on GD 7 (200 mg/kg bw/d) or GD 18 (60 mg/kg bw/d). Salivation occurred
most probably due to the bad taste of the test substance, was assessed to be a local effect
and, therefore, considered as not adverse.
No clinical signs or changes of general behavior, which may be attributed to the test substance,
were detected in any female of test group 1 (20 mg/kg bw/d).
see attached document: summary of maternal clinical observation

Description (incidence):

Due to severe clinical findings in almost all animals of test group 3, the high-dose group of 200 mg/kg bw/d was sacrificed prematurely due to animal
welfare reasons on GD 13/12/11 ) of the study.
There were no further test substance-related or spontaneous mortalities in any females of test
groups 0-2 (0, 20 or 60 mg/kg bw/d).

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

see attached document: summary bw_bwc
From GD 8 onwards until premature sacrifice on GD 11-13 the mean body weights of the highdose
dams (200 mg/kg bw/d) were lower in comparison to the concurrent control group,
attaining statistical significance on GD 10 (-4%). The average body weight gain of the highdose
dams (200 mg/kg bw/d) was distinctly and statistically significantly reduced during GD 6-
10 showing a body weight loss during GD 6-8. These effects were assessed as
treatmentrelated and adverse.
In test group 2 (60 mg/kg bw/d) the mean body weights were generally comparable to the
concurrent control group. The average body weight gain was statistically significantly reduced
on GD 6-8 but recovered afterwards and was comparable to the control values again
throughout the remaining study period. If calculated for the entire treatment period (GD 6-19),
the mean body weight gain of the mid-dose dams was comparable to the concurrent control
group.
The mean body weights and the average body weight gain of the low-dose dams (20 mg/kg
bw/d) were generally comparable to the concurrent control group throughout the entire study
period.
Corrected (net) body weight gain
The corrected body weight gain of test groups 1 and 2 (20 and 60 mg/kg bw/d) revealed no
difference of any biological relevance to the corresponding control group. Moreover, mean
carcass weights remained unaffected by the treatment.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

see attached document: summary food consumption
The mean food consumption of the high-dose dams (200 mg/kg bw/d) was statistically
significantly reduced from GD 6 onwards until premature sacrifice on GD 11-13 (up to 31%
below control). This was assessed as treatment-related and adverse.
The mean food consumption of the mid- and low-dose dams (60 and 20 mg/kg bw/d) was
generally comparable to the concurrent control group throughout the entire study period.

Water consumption and compound intake (if drinking water study):

effects observed, treatment-related

Description (incidence and severity):

see attached document; summary maternal water consumption
The mean water consumption of the dams in test group 3 (200 mg/kg bw/d) was statistically
significantly reduced from GD 6 onwards until premature sacrifice during GD 11-13 (up to 21%
in comparison to the concurrent control). This was assessed as treatment-related and adverse.
In test group 2 (60 mg/kg bw/d) the mean water consumption was statistically significantly
reduced during GD 6-10 (up to 12% below control) but recovered afterwards and was
comparable to the control values again until scheduled sacrifice on GD 20.
The mean water consumption of the dams in test group 1 (20 mg/kg bw/d) was comparable to
the concurrent control group throughout the entire study period.

Ophthalmological findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Description (incidence and severity):

see attached document: summary gravid uterine weitghts and bwc
The mean gravid uterus weights of the low- and mid-dose animals (20 and 60 mg/kg bw/d)
were not influenced by the test substance. The differences between these groups and the
control group revealed no dose-dependency and were assessed to be without biological
relevance.
Absolute and relative organ weights
All mean absolute and relative weight parameters did not show significant differences when
compared to the control group 0.

Histopathological findings: non-neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

All findings occurred either individually or were biologically equally distributed over control and
treatment groups. They were considered to be incidental or spontaneous in origin and without
any relation to treatment.

Other effects:

no effects observed

Description (incidence and severity):

See attached document: Incidence of microscopic findings
Thyroid hormones
At gestation day 20, in dams of test groups 1 and 2 (20 and 60 mg/kg bw/d) no treatment related
alterations of T3, T4 and TSH were observed.
Gross lesions
No macroscopic findings of thyroid glands were noted in test groups 0 (control), 1 (20 mg/kg
bw/d) and 2 (60 mg/kg bw/d).

Maternal developmental toxicity

Number of abortions:

no effects observed

Pre- and post-implantation loss:

effects observed, non-treatment-related

Description (incidence and severity):

A 100% post-implantation loss was observed in one female each in test groups 1 and 2. In one lowdose
dam (20 mg/kg bw/d) one early resorption was recorded after staining the empty
uterus at C-section (i.e. pregnant by stain), whereas for one mid-dose dam (60 mg/kg bw/d)
9 early and 3 late resorptions were recorded (no viable fetuses). In the other dams of test
groups 1 and 2 post-implantation losses and number of resorptions were comparable to the
control. Since only one dam per test group was affected without relation to dose, these isolated
findings were regarded as incidental and not treatment-related.

Total litter losses by resorption:

no effects observed

Description (incidence and severity):

see attached document: summary of reproduction data
There were no test substance-related and/or biologically relevant differences between the test
groups 0-2 in conception rate, in the mean number of corpora lutea and implantation sites or
in the values calculated for the pre- and post-implantation losses, the number of resorptions
and viable fetuses. All observed differences are considered to reflect the normal range of
fluctuations for animals of this strain and age.

Early or late resorptions:

effects observed, non-treatment-related

Description (incidence and severity):

In one lowdose dam (20 mg/kg bw/d) one early resorption was recorded after staining the empty
uterus at C-section (i.e. pregnant by stain), whereas for one dam in the mid-dose (60 mg/kg bw/d)
9 early and 3 late resorptions were recorded (no viable fetuses).

Dead fetuses:

no effects observed

Changes in pregnancy duration:

no effects observed

Changes in number of pregnant:

no effects observed

Details on maternal toxic effects:

There were no test substance-related and/or biologically relevant differences between the test
groups 0-2 in conception rate, in the mean number of corpora lutea and implantation sites or
in the values calculated for the pre- and post-implantation losses, the number of resorptions
and viable fetuses. All observed differences are considered to reflect the normal range of
fluctuations for animals of this strain and age.
Regarding pathology, no treatment-related findings were noted. All findings occurred either
individually or were biologically equally distributed over control and treatment groups. They
were considered to be incidental or spontaneous in origin and without any relation to treatment.

Effect levels (maternal animals)

Key result

Dose descriptor:

NOAEL

Effect level:

60 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

body weight and weight gain

clinical signs

food consumption and compound intake

water consumption and compound intake

Results (fetuses)

Fetal body weight changes:

no effects observed

Description (incidence and severity):

see attached document: summary placental and fetal bw
The mean fetal weights of test groups 1 and 2 (20 and 60 mg/kg bw/d) were not influenced by
the test substance and did not show any biologically relevant differences in comparison to the
control group.

Changes in sex ratio:

no effects observed

Description (incidence and severity):

The sex distribution of the fetuses in test groups 1 and 2 (20 and 60 mg/kg bw/d) was
comparable to the control fetuses.

Changes in litter size and weights:

no effects observed

Changes in postnatal survival:

not examined

External malformations:

effects observed, non-treatment-related

Description (incidence and severity):

One low-dose male fetus an umbilical hernia was recordrd. Since the finding was not related to dose,
it was not assessed as treatment-related.

Skeletal malformations:

effects observed, non-treatment-related

Description (incidence and severity):

For only one low-dose fetus a skeletal malformation was recorded: malpositioned and bipartite
sternebra (unchanged cartilage). Since the finding was not related to dose, it was not assessed
as treatment-related.

Visceral malformations:

effects observed, non-treatment-related

Description (incidence and severity):

Two soft tissue variations were detected, i.e. short innominate and dilated renal pelvis, both in
test groups 0 and 1. The incidences of these variations were neither statistically significantly
nor dose-dependently increased in the treated groups. Therefore, they were not assessed as
treatment-related.

Other effects:

effects observed, non-treatment-related

Description (incidence and severity):

Two unclassified external observations were recorded. A skin tag was seen in one low-dose
fetus (20 mg/kg bw/d), furthermore, placentae fused were seen in one control litter. These
findings were not considered biologically relevant, since they were single events without
relation to dose.
For all test groups, skeletal variations of different bone structures were observed, with or
without effects on corresponding cartilages. The observed skeletal variations were related to
several parts of fetal skeletons and appeared without a relation to dose The overall affected
fetuses/litter incidences of skeletal variations were comparable to the historical control data.

Details on embryotoxic / teratogenic effects:

Soft tissue malformations did not occur in any fetus in this study. There were noted external
and skeletal malformations in two fetuses of the low-dose group.. These observations in single
fetuses were unrelated to dose and both can be found in the historical control data, thus they
are considered as incidental events.
External variations did not occur in any fetus in this study. Two soft tissue variations and a range of
skeletal variations were noted in all test groups including the controls. Two out of three incidences
showed no relation to dosing. The skeletal variations are equally distributed about the different
test groups, if normal biological variation is taken into account, and can be found in the historical
control data at a comparable frequency.
Unclassified soft tissue observations did not occur in any of the fetuses in this study. A
spontaneous origin is assumed for the unclassified external and skeletal cartilage observations
which were observed in several fetuses of all test groups The distribution and type of these findings
do not suggest any relation to treatment.

Effect levels (fetuses)

Key result

Dose descriptor:

NOAEL

Effect level:

60 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: teratogenicity / embryotoxicity

Fetal abnormalities

Key result

Abnormalities:

no effects observed

Overall developmental toxicity

Key result

Developmental effects observed:

no

Treatment related:

no

Any other information on results incl. tables

Total soft tissue variations

 

		Test group 0 0 mg/kg bw/d	Test group 1 20 mg/kg bw/d	Test group 2 60 mg/kg bw/d
Litter Fetuses	N N	25 133	24 121	24 121
Fetal incidence	N (%)	3 (2.3)	3 (2.5)	0.0
Litter incidence	N (%)	3 (12)	2 (8.3)	0.0
Affected fetuses/litter	Mean %	2.0	2.5	0.0

 

mg/kg bw/d = milligram per kilogram body weight per day; N = number; % = per cent

Total fetal skeletal variations

 

		Test group 0 0 mg/kg bw/d	Test group 1 20 mg/kg bw/d	Test group 2 60 mg/kg bw/d
Litter Fetuses	N N	25 144	24 129	24 132
Fetal incidence	N (%)	138 (96)	127 (98)	129 (98)
Litter incidence	N (%)	25 (100)	24 (100)	24 (100)
Affected fetuses/litter	Mean %	95.9	98.6	97.9

 

mg/kg bw/d = milligram per kilogram body weight per day; N = number; % = per cent

 

 

 

Occurrence of statistically significantly increased fetal skeletal         

variations (expressed as mean percentage of affected fetuses/litter)

 

Finding	Test group 0 0 mg/kg bw/d	Test group 1 20 mg/kg bw/d	Test group 2 60 mg/kg bw/d	HCD Mean % (range)
Incomplete ossification of basisphenoid	10.4	32.8**	18.4	22.4 (5.3 - 43.2)
Incomplete ossification of supraoccipital; unchanged cartilage	11.8	17.2	23.6*	22.4 (3.0 - 51.2)
Incomplete ossification of skull; unchanged cartilage	3.5	9.2*	5.0	7.5 (2.1 - 13.4)

 

mg/kg bw/d = milligram per kilogram body weight per day; HCD = Historical control data; % = per cent

* = p≤0.05 (Wilcoxon-test [one-sided]) ** = p≤0.01 (Wilcoxon-test [one-sided])

 

Applicant's summary and conclusion

Conclusions:

Under the conditions of this prenatal developmental toxicity study, the oral administration of
Dibutylethanolamine to pregnant Wistar rats from implantation to one day prior to the
expected day of parturition (GD 6-19) caused evidence of distinct maternal toxicity, such as
severe clinical findings (twitching, convulsions, tremor, abdominal position), a reduction of
water and food consumption and a decrease in body weight/body weight gain. Due to the
above-mentioned findings, treatment of the high-dose was stopped and the dams were
euthanized without further examinations on GD 13/12/11 of the study.
In conclusion, the no observed adverse effect level (NOAEL) for maternal toxicity is the
mid-dose of 60 mg/kg bw/d.
Examination of the low- and mid-dose fetuses was performed and showed no adverse findings.
In conclusion, the no observed adverse effect level (NOAEL) for prenatal developmental
toxicity is 60 mg/kg bw/d.
Under the conditions of this study the test item is not teratogenic.

Executive summary:

In a prenatal developmental toxicity study, the test substance Dibutylethanolamine was

administered to pregnant Wistar rats daily by gavage from implantation to one day prior to the

expected day of parturition (GD 6-19) to evaluate its potential maternal and prenatal

developmental toxicity.

Analyses confirmed the correctness of the prepared concentrations and the stability of the test

substance in the vehicle.

Concerning clinical examination, strong signs of systemic maternal toxicity was observed at

the highest test group of 200 mg/kg bw/d Dibutylethanolamine. Since severe clinical findings

(twitching, convulsions, tremor, abdominal position) were observed in almost all animals, the

high-dose group was sacrificed prematurely due to animal welfare reasons. Treatment of that

test group was stopped and the dams were euthanized without further examinations on GD

13/12/11 of the study. The next lower dose of 60 mg/kg bw/d showed no adverse clinical

findings.

Water and food consumptions were statistically significantly reduced in high-dose dams during

GD 6-13 (up to 21% and 31% below control, respectively). Furthermore, high-dose dams

showed a statistically significant decrease in body weight (change) including a body weight

loss during GD 6-8. The above-mentioned findings were assessed as treatment-related and

adverse.

Mid-dose dams showed a reduction in water consumption in the beginning of treatment (GD

6-10: up to 12% below control) but recovered afterwards to control levels. Food consumption

was not affected in mid-dose dams. Body weight change was statistically significantly reduced

on GD 6-8 but recovered afterwards and body weight in general was comparable to the control

values. These two marginal findings in the mid-dose group were only observed in the beginning

of treatment but not thereafter. They were considered to be not sufficient to proof adversity and

therefore, they were not assessed as treatment-related and adverse.

In the low dose group (20 mg/kg bw/d), no adverse findings were observed.

Concerning thyroid hormone levels, no treatment-related, adverse effect was observed up to a

dose of the compound of 60 mg/kg bw/d.

Regarding pathology, no treatment-related findings were noted. All findings occurred either

individually or were biologically equally distributed over control and treatment groups. They

were considered to be incidental or spontaneous in origin and without any relation to treatment.

No differences of toxicological relevance between the control and the treated groups (20 or 60

mg/kg bw/d) were determined for any reproductive parameters, such as conception rate, mean

number of corpora lutea, mean number of implantations, as well as pre- and post-implantation

loss. Similarly, no toxicologically relevant influence of the test substance on sex distribution

and anogenital distance/index of the fetuses was noted at any dose.

Overall, there was no evidence for toxicologically relevant adverse effects of the test substance

on fetal morphology at any dose.