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Toxicological information

Developmental toxicity / teratogenicity

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Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
13 October 2020 - 04 January 2021.
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2021
Report date:
2021

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
July 2018
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
1-(N,N-bis(2-hydroxyethyl)amino)propan-2-ol
EC Number:
229-764-5
EC Name:
1-(N,N-bis(2-hydroxyethyl)amino)propan-2-ol
Cas Number:
6712-98-7
Molecular formula:
C7H17NO3
IUPAC Name:
1-[bis(2-hydroxyethyl)amino]propan-2-ol
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source: Sasol, Marl, Germany; Batch no. 200162527.
- Purity: 99.6%.
- Appearance: clear liquid.

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: At ambient temperature, in the dark.

Test animals

Species:
rabbit
Strain:
New Zealand White
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Envigo RMS, UK.
- Age at study initiation: 17 - 22 weeks.
- Weight at study initiation: 2.30 - 4.17 kg.
- Fasting period before study: No.
- Housing: One animal (female) per cage.
- Diet: Commercial diet, 200 g/animal/day. "A small supplement of autoclaved hay was given on a daily basis to promote gastric motility and a small amount of chopped fresh vegetables were given twice weekly."
- Water: Tap water ad libitum.
- Acclimation period: Five days before commencement of treatment (days 1 - 5 after mating).

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 15 - 21 °C.
- Humidity (%): 45 - 70%.
- Air changes (per hr): Not specified. Filtered fresh air which was passed to atmosphere and not recirculated.
- Photoperiod: 14 hours light / 10 hours dark.

IN-LIFE DATES:
From: Approx. July 2020 (based on age on arrival at testing facility).
To: 24 - 27 November 2020.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Water was added to pre-weighed test material and mixed (for at least 40 minutes) to obtain a homogenous solution at the highest concentration. Lower concentrations were prepared by serial dilution.

VEHICLE
- Concentration in vehicle: 25, 75 and 250 mg/mL
- Amount of vehicle (if gavage): 4 mL/kg bw.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples of each formulation prepared for administration in the first and last preparation were analyzed for achieved concentration of the test item.
Details on mating procedure:
Animals were received at the test facility having already been successfully mated. Natural mating with New Zealand white bucks of established fertility at the supplier’s facility. Males and females were not closely related.
Duration of treatment / exposure:
Days 6 - 28 (inclusive) after mating (i.e. 23 days total).
Frequency of treatment:
Once daily.
Doses / concentrationsopen allclose all
Dose / conc.:
100 mg/kg bw/day (nominal)
Dose / conc.:
300 mg/kg bw/day (nominal)
Dose / conc.:
1 000 mg/kg bw/day (nominal)
No. of animals per sex per dose:
24
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Dose levels selected for this study, were based on the results from a Combined Pilot and Preliminary Study.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes.
- Time schedule: At least twice daily.
- Specific cage side observations not listed in study report.

DETAILED CLINICAL OBSERVATIONS: Yes.
- Time schedule: At least twice daily.

BODY WEIGHT: Yes.
- Time schedule for examinations: On arrival (day 1 after mating), day 3 after mating, and daily on days 6 - 29 after mating.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on day 29 after mating.
- Organs examined: A full macroscopic examination of the tissues was performed. All external features and orifices were examined visually. Any abnormality in the appearance or size of any organ and tissue (external and cut surface) was recorded and the required tissue samples preserved in appropriate fixative.
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes.
Examinations included:
- Gravid uterus weight: Yes (including cervix and ovaries).
- Number of corpora lutea: Yes.
- Number of implantations: Yes.
- Number of early resorptions: Yes.
- Number of late resorptions: Yes.
- Other: Number of foetuses (live and dead).
Fetal examinations:
- External examinations: Yes [all per litter].
- Soft tissue examinations: Yes [all per litter].
- Skeletal examinations: Yes [all per litter].
- Head examinations: Yes [50% per litter].
- Anogenital distance of all live rodent pups: Not applicable.
Statistics:
The following data types were analyzed at each timepoint separately:
Body weight, using absolute weights and gains over appropriate study periods
Gravid uterine weight and adjusted body weight
Food consumption, over appropriate study periods
C-section litter data (corpora lutea, implantations, pre/post implantation loss, live young and sex ratio - percentage male)
Placental, litter and fetal weights

A parametric analysis, using the t-test or F1 approximate test, was performed if Bartlett's test was not significant at the 1% level.

A non-parametric analysis, using Kruskal-Wallis' test, Wilcoxon rank sum tests, or the H1 approximate test, was performed if Bartlett's test was still significant at the 1% level following both logarithmic and square-root transformations.
Historical control data:
Historical control data from 14 studies conducted from January 2020 are included, using New Zealand white rabbits from the same supplier (Envigo RMS) as used in the present study.

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
no effects observed
Mortality:
mortality observed, non-treatment-related
Description (incidence):
One animal in each of the vehicle control, mid-dose and high-dose groups was killed early for welfare reasons.

In the case of the control and mid-dose animals, the litters had resorbed. "The poor clinical condition of [the mid-dose animal] was attributed to the loss of the litter and not attributable to treatment with DEIPA."

The high-dose animal was killed for welfare reasons on suspicion that the pregnancy had resorbed or aborted. Upon examination, the death of the animal was due to a twisted caecum, and not attributable to treatment with DEIPA.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
See Table 4.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Food consumption was 18% lower than control in the high-dose group. See Table 1.
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
Organ weight findings including organ / body weight ratios:
not specified
Gross pathological findings:
no effects observed
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined

Maternal developmental toxicity

Number of abortions:
not specified
Description (incidence and severity):
Data not available in study report.
Pre- and post-implantation loss:
no effects observed
Description (incidence and severity):
See Table 3.
Total litter losses by resorption:
no effects observed
Early or late resorptions:
no effects observed
Dead fetuses:
not specified
Description (incidence and severity):
Data on stillbirths and/or dead foetuses not available in study report.
Changes in pregnancy duration:
not specified
Description (incidence and severity):
Data, including on early deliveries, not available in study report.
Changes in number of pregnant:
no effects observed
Description (incidence and severity):
11 animals were not pregnant: 3 in the control group, 2 in the low-dose group, 4 in the mid-dose group and 2 in the high-dose group. See Table 2.

Effect levels (maternal animals)

Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Remarks on result:
not determinable due to absence of adverse toxic effects

Results (fetuses)

Fetal body weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
Total litter weight was marginally reduced in the high-dose group, attributed to the marginally smaller litter size.
Reduction in number of live offspring:
effects observed, non-treatment-related
Description (incidence and severity):
"Marginally smaller" litter size at the high dose. See Table 5.
Changes in sex ratio:
no effects observed
Description (incidence and severity):
See Table 5.
Changes in litter size and weights:
no effects observed
Description (incidence and severity):
See Table 6.
Changes in postnatal survival:
not examined
External malformations:
no effects observed
Description (incidence and severity):
See Table 7.
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
There was 1 incidence each of lumbar scoliosis, lumbar hemivertebra, bent clavicle and pointing backwards hindpaw(s). See Table 7.

Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
There were 2 incidences each of dilated ascending aorta and narrow pulmonary trunk, and 1 each of acephalostomia, truncus arteriosus and septal defect. See Table 7.
Other effects:
effects observed, non-treatment-related
Description (incidence and severity):
The incidence of incompletely ossified/unossified epiphyses and metacarpals/phalanges at all treated doses exceeded the concurrent and historical controls ranges. These are a transient stage in foetal development and indicative of foetal immaturity, and therefore non-adverse. See Table 8.

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects

Overall developmental toxicity

Key result
Developmental effects observed:
no

Any other information on results incl. tables

Table 1 - Food consumption during gestation days 6-29



































GroupNominal dose (mg/kg bw/day)Mean food consumption (g/animal/day)Standard Deviation
1010214.6
210010011.7
33009714.0
4100084**14.7

** p < 0.01


 


Table 2 - Number of pregnancies













































GroupNominal dose (mg/kg bw/day)Number of treated animalsNumber of animals that died during studyNumber of non-pregnant damsNumber of pregnant dams
10241320
2100240222
3300241419
41000241221

 


Table 3 - Pre- and post-implantation loss - litter data






















































































GroupNominal dose (mg/kg bw/day)Number of pregnant damsResorptionsPre-implantation loss (%)Post-implantation loss (%)
EarlyLateTotal
10200.60.10.710.08.7
SD 0.94SD 0.31SD 0.92SD 12.08SD 11.87
2100220.70.41.015.312.9
SD 1.36SD 0.49SD 1.36SD 22.51SD 15.66
3300190.60.40.98.810.3
SD 1.02SD 0.76SD 1.18SD 12.23SD 12.05
41000210.50.30.813.310.8
SD 0.98SD 0.64SD 1.21SD 18.31SD 16.00

 


Table 4 - Body and gravid uterine weights




























































































  Group mean values (kg)
GroupNominal dose (mg/kg bw/day)Body weight, day 6Terminal body weightBody weight changeGravid uterine weightAdjusted terminal body weightAdjusted body weight change
103.133.420.290.4203.00-0.13
SD 0.415SD 0.343SD 0.133SD 0.0997SD 0.343SD 0.154
21003.193.450.250.3963.05-0.14
SD 0.374SD 0.318SD 0.145SD 0.1325SD 0.222SD 0.175
33003.133.410.280.4083.00-0.13
SD 0.321SD 0.301SD 0.115SD 0.0718SD 0.278SD 0.102
410003.083.360.280.3662.99-0.08
SD 0.323SD 0.307SD 0.174SD 0.0990SD 0.274SD 0.132

 


Table 5 - Live offspring








































































GroupNominal dose (mg/kg bw/day)Mean number of live youngSex ratio (% male)
MaleFemaleTotal
103.93.67.550.3
SD 2.13SD 1.27SD 2.37SD 14.27
21004.22.87.062.8
SD 2.18SD 2.14SD 2.60SD 25.75
33003.14.27.341.5
SD 1.66SD 1.65SD 1.53SD 21.33
410003.03.76.743.7
SD 1.70SD 1.76SD 2.19SD 19.50

 


Table 6 - Litter and foetal weights
















































































GroupNominal dose (mg/kg bw/day)Mean total litter weight (g)Mean foetal weights (g)
MaleFemaleCombined
10279.539.237.538.4
  SD 71.98SD 6.26SD 5.97SD 5.68
2100260.638.036.738.0
  SD 96.02SD 6.86SD 6.03SD 5.99
3300268.436.637.337.2
  SD 55.61SD 4.93SD 4.68SD 4.71
41000242.936.236.236.6
  SD 74.07SD 6.06SD 4.80SD 4.40

 


Table 7 - Foetal examinations - major abnormality findings


































































































































































Group1234
Nominal dose (mg/kg bw/day)01003001000
Litters examined20221921
Total number of litters affected1002
Foetuses examined150154139141
Total number of foetuses affected3004
Number of foetuses with major abnormalities
SkeletalHeadAcephalostomia0001
Cervical/ThoracicThoracic scoliosis1000
Dorsoventral distortion of sternum1000
Lubmar (and abdominal)/ sacral/ caudalSacral/caudal spina bifida1000
Lumbar scoliosis0001
Lumbar hemivertebra0001
AppendicularBent clavicle0001
Pointing backwards hindpaw0001
VisceralCervical/ThoracicDilated ascending aorta/aortic arch0002
Dorsally displaced pulmonary trunk1000
Truncus arteriosus0001
Membranous ventricular septal defect1001
Muscular ventricular septal defect0001
Fistula ascending aorta/pulmonary trunk1000
Narrow/marked pulmonary trunk0002

 


Table 8 - Foetal examinations - minor abnormality findings


































































































































































































































































































































































































































Group1234
Nominal dose (mg/kg bw/day)01003001000
Litters examined20221921
Foetuses examined150154139141
Number of foetuses with minor abnormalities
SkeletalCranial abnormality - unossified area(s)0010
Cranial abnormality - sutural bone(s)1000
Cranial abnormality - bent cornu(a) of hyoid0833
Vertebral abnormality - thoracic1001
Vertebral abnormality - lumbar1001
Rib abnormality - short without costal cartilage1000
Rib abnormality - misaligned1000
Rib abnormality - partially fused1000
Rib abnormality - distally thickened1000
Rib abnormality - branched with additional costal cartilage1000
Sternebrae - fused/partially fused3314
Sternebrae - misaligned hemicentres0210
Sternebrae - supernumerary site0100
Sternebrae - wide1000
Sternebrae - misshapen2000
Sternebrae - branched 6th0100
Costal cartilage - partially fused2000
Costal cartilage - misaligned0312
Costal cartilage - branched1100
Costal cartilage - additional1000
Costal cartilage - 7th not connected to sternum0200
Appendicular abnormality - absent 1st digit forepaw(s)0001
Cervical rib abnormality - short supernumerary1972
Cervical rib abnormality - full supernumerary1000
Number of 13th ribs - short supernumerary44343745
Number of 13th ribs - full supernumerary35322257
Number of 13th ribs - total67615489
Thoracolumbar vertebrae - 181010
Thoracolumbar vertebrae - 20712419
Pelvic girdle - unilateral cranial shift2000
Pelvic girdle - unilateral caudal shift6204
Delayed/incomplete ossificationCranial - large posterior fontanelle1001
Sternebrae - 5th20233924
Sternebrae - 1st-4th, 6th951113
Sternebrae - total28264632
Vertebrae - cervical3002
Vertebrae - thoracic1000
Appendicular - pubes0201
Appendicular - epiphyses11201014
Appendicular - talus1100
Appendicular - metacarpals/phalanges9161517
Appendicular - metatarsals/phalanges2714
Increased ossificationCranial - partially fused jugal to maxilla1100
Vertebrae - long lumbar transverse processes1000
VisceralEyes - folded retina3200
Brain - dilated interventricular foramen1000
Head - supernumerary minor upper incisor0100
Lungs - abnormal lobation1000
Gall bladder - bilobed0010
Ovary - cyst(s)0010
Head - tongue present0001
Limb(s) - flexure forepaw(s)1103
Tail - curled0001
Tail - kinked0200

Applicant's summary and conclusion

Conclusions:
In a good-quality guideline study to GLP, gavage administration of DEIPA at doses up to 1000 mg/kg bw/day (nominal) to time-mated rabbits on GD 6-28 (inclusive) did not produce any clinical signs of toxicity. Reproductive parameters were not altered and no signs of treatment-related malformations were seen. On this basis, the study NOAELs for both maternal and developmental toxicity were 1000 mg/kg bw/day (nominal), the highest dose tested.
Executive summary:

In a good quality guideline study, conducted to GLP, groups of 24 time-mated female New Zealand white rabbits were administered DEIPA by gavage at dose levels of 0, 100, 300, or 1000 mg/kg bw/day (nominal) on days 6-28 of gestation. Animals were killed on gestation day 29 and examined for gross pathological alterations. Gravid uterine weights were recorded, along with the number of corpora lutea, implantations, resorptions, and live/dead foetuses. All foetuses were weighed and sexed, and examined for external, visceral and (in 50%) skeletal alterations.


 


No clinical signs of toxicity were seen, reproductive parameters were not altered, and there were no treatment-related malformations or other developmental toxicity observed in the foetuses at any dose of DEIPA. On this basis, the no-observed-adverse-effect levels (NOAELs) for maternal toxicity and developmental toxicity were 1000 mg/kg bw/day (nominal), the highest dose tested.

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