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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:
Sep - Nov 1983
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
comparable to guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1984
Report date:
1984

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Ammonium 2-amino-4-(hydroxymethylphosphinyl)butyrate
EC Number:
278-636-5
EC Name:
Ammonium 2-amino-4-(hydroxymethylphosphinyl)butyrate
Cas Number:
77182-82-2
Molecular formula:
C5H12NO4P.H3N
IUPAC Name:
ammonium 2-amino-4-(hydroxymethylphosphinyl)butyrate

Test animals

Species:
rabbit
Strain:
Himalayan
Remarks:
Hoe:HIMK(SPFWiga)
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Hoechst AG
- Age at study initiation: 6-7 months
- Weight at study initiation: 2547 +/- 203 g
- Fasting period before study:
- Housing: individually
- Diet: ad libitum plus 40-50 g autoclaved hay
- Water: ad libitum
- Acclimation period: not specified

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22
- Humidity (%): 47-65
- Air changes (per hr): 16-20
- Photoperiod (hrs dark / hrs light): 10/14, illumination from 6 am to 8 pm

IN-LIFE DATES: From: To:

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Remarks:
distilled water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: daily
- an equal liquid volume of 5 ml/kg bodyweight was administered to each animal between 8 and 11 a.m.
Analytical verification of doses or concentrations:
no
Details on mating procedure:
- Impregnation procedure: [artificial insemination / purchased timed pregnant / cohoused]
- If cohoused: yes
- M/F ratio per cage: not specified
- Length of cohabitation: In the morning hours during estrous
- When sperm was detected, animals were paired again 6 h later
- Determination of GD: Day of pairing was counted as day 0 of gravidity
- Proof of pregnancy: presense of implantation sites in the uterus
Duration of treatment / exposure:
GD 7-19
Frequency of treatment:
once daily
Duration of test:
On GD 29, the dams were killed and delivered.
Doses / concentrationsopen allclose all
Dose / conc.:
2 mg/kg bw/day
Dose / conc.:
6.3 mg/kg bw/day
Dose / conc.:
20 mg/kg bw/day
No. of animals per sex per dose:
15
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Before the start of the study, the test substance was administered in a range-finding test at the dose levels 10.0, 22.4 and 50.0 mg/kg bodyweight, in each case to 2 gravid Himalayan rabbits. In this test, 10.0 mg/kg was tolerated without complications by dams and foetuses alike. At 22.4 mg/kg bodyweight, both dams showed reduced body weights and one showed, in addition to six normally developed live foetuses, three dead foetuses and a conceptus under resorption in the uterus. At 50.0 mg/kg, a decrease in body weight, food consumption, and marked clinical signs of intolerance were observed. One of the animals was therefore killed on the following day (day 14 of gravidity), and the other died in the night from days 15 to 16 of gravidity. Based on these results, the following dose levels were established for the main study:
2.0 mg/kg as the dose likely to be tolerated by both dams and foetuses;
6.3 mg/kg as the geometrical mean between the selected low and high doses;
20.0 mg/kg as the dose likely to produce signs of intolerance.

- Rationale for animal assignment (if not random): random, scheme No. 364/83

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: weekly in the first three weeks and then once again after a further 9 days.

FOOD CONSUMPTION: Yes
- Time schedule: continuous

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 29
- Organs examined: uterus, heart, liver, kidneys, spleen
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: No
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of resorptions: Yes
- Other: conceptuses, placentae, placental weights
Fetal examinations:
- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes: half per litter
- Head examinations: No
Statistics:
Dunnett, Nemenzi/Dunnett, Goodman, Wilcoxon (pairwise)

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
A decrease in water consumption and/or quantity of faeces was observed between GD13 and 25 in 4 dams of the 2.0 mg/kg group, 2 dams of the 6.3 mg/kg group, 7 dams of the 20.0 mg/kg group and one control dam. The same findings were observed in the dams which did not survive until the end of the study.
Dermal irritation (if dermal study):
no effects observed
Mortality:
mortality observed, treatment-related
Description (incidence):
One dam from the 6.3 mg/kg/day treated group died on GD29 while giving premature birth. One dam from the 20 mg/kg/day treated group was sacrificed on GD17 after having shown extension spasms for 5 seconds immediately after administration on GD16 followed by high-legged position with the head stretched out and tilted for 10 minutes and then a phase of head tilting and drowsiness. On GD17 just before sacrifice the animal was lying on its stomach in a state of apathy. Another dam from this group aborted in the night from GD19 to 20 and a third one delivered prematurely in the night from GD24 to 25.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
- 20 mg/kg bw/day: decreased body weights during the first week of treatment and continued thereafter until GD 29
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
- 6.3 mg/kg bw/day: slight decrease in food consumption during the first week of treatment. However, this had no effect on body weight gains and was therefore regarded as a not particulary important finding.
- 20 mg/kg bw/day: marked decrease of food consumption in the first week of treatment and a moderate decrease in the second week.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
effects observed, treatment-related
Description (incidence and severity):
- 2.0 mg/kg bw/day: decrease in water consumption in four dams
- 6.3 mg/kg bw/day: decrease in water consumption in 2 dams
- 20 mg/kg bw/day: decrease in water consumption in 7 dams
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
effects observed, treatment-related
Description (incidence and severity):
- 20 mg/kg bw/day: one dam showed extension spasms (ca. 5 sec) immediately after the 10th application on GD 16; the animal subsequently remained for about 10 minutes in high-legged position, with the head stretched out and tilted, followed by a phase of head tilting and drowsiness; on the next morning the dam was lying on its stomach in a state of apathy
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
- 20 mg/kg bw/day: Kidney weights were slightly higher. All dams showed relatively high liver weights (not statistically significant and within the range of historical controls).
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
- 6.3 mg/kg bw/day: dark brown areas from 0.3 to 0.5 mm in diameter on the surface of both kidneys in one dam.
- 20 mg/kg bw/day: a cyst approximately 3 mm in size with greenish brown liquid contents was found on the surface of the lobus dexter accessorius of the liver in one dam, and also on the surface of the lobus caudatus of the liver in another dam. One dam showed recesses of different sizes on the surface of both kidneys.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined

Maternal developmental toxicity

Number of abortions:
effects observed, treatment-related
Description (incidence and severity):
-20 mg/kg bw/day: increase in the number of abortions
Pre- and post-implantation loss:
no effects observed
Total litter losses by resorption:
no effects observed
Early or late resorptions:
effects observed, non-treatment-related
Description (incidence and severity):
The total number of resorptions in the treatment groups was in the same range as that in the control group.
Dead fetuses:
effects observed, non-treatment-related
Description (incidence and severity):
- 2.0 mg/kg bw/day: all 15 dams bore live fetuses. Three dams bore one dead fetus each.
- 6.3 mg/kg bw/day: 14 dams bore live fetuses. One dam presented 5 plancentae and 5 living and 3 dead fetuses, another 8 living and one dead fetus
- 20 mg/kg bw/day: 11 dams bore live fetuses. One had seven implantation sites and two conseptuses under resorption in the uterus, but no living fetuses. One dam presented 6 dead fetuses, another revealed 7 conceptuses and a resorption site, and the dam that aborted bore 6 live fetuses and one dead fetus. From the dams that bore live fetuses, four bore one dead fetus and one bore two dead fetuses.

The total number of dead foetuses and resorptions in the treatment groups was in the same range as that in the control group.
Changes in pregnancy duration:
effects observed, treatment-related
Description (incidence and severity):
- 20 mg/kg bw/day: increase in premature births
Changes in number of pregnant:
no effects observed
Other effects:
no effects observed
Description (incidence and severity):
The number of corpora lutea, as an indication of the number of ovulations which had occurred, and the number of implantations were within the same range for the treated dams as for the controls.

Effect levels (maternal animals)

Dose descriptor:
NOAEL
Effect level:
6.3 mg/kg bw/day
Based on:
act. ingr.
Basis for effect level:
body weight and weight gain
clinical signs
food consumption and compound intake
number of abortions
organ weights and organ / body weight ratios
water consumption and compound intake
other: increase in premature births

Maternal abnormalities

Abnormalities:
effects observed, treatment-related
Localisation:
uterus
Description (incidence and severity):
An increase in the number of premature births and abortions was observed at 20 mg/kg bw/day.

Results (fetuses)

Fetal body weight changes:
no effects observed
Reduction in number of live offspring:
effects observed, non-treatment-related
Description (incidence and severity):
The live foetuses delivered in all three treatment groups were normally developed. The total number of dead foetuses in the treatment groups was in the same range as that in the control group.
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
no effects observed
Description (incidence and severity):
No disturbance of the viability of the foetuses during the first 24 hours after delivery.
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
- 2 mg/kg bw/day: One fetus showed an encephalocele which was due to a marked fissuration in the region of the os frontale and os parietale. This foetus died after one hour in the incubator.
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
The skeletons of the live foetuses in the treatment groups were at the same stage of development as the control foetuses. Skeletons of the foetuses found dead in the uterus were more weakly ossified.
- 2 mg/kg bw/day: calvarial fissuration in one fetus. One fetus showed fragmentation of the right dorsal vertebral arch on the 2nd vertebra.
- 6.3 mg/kg bw/day: slight splintering on the os parietale in one fetus. One or more fetuses showed a rib primordium on the 7th cervical vertebra
- 20 mg/kg bw/day: slight splintering on the os parietale in one fetus and another showed fused caudal vertebrae.

Other foetuses in all groups a primordium of a 13th rib or fused, dysplastic and dislocated sternebrae.
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Fetuses in all groups revealed aplasia of the lobus inferior medialis of the lung and/or adhesion of pulmonary lobes. One or more foetuses in all
groups also showed the stomach in transverse position, or taut with fluid and/or enlarged. Caudad displacement and/or transverse position of the left kidney was also encountered in a number of foetuses. Blood was found in the pericardium in one foetus of the 2.0 mg/kg group, and in the abdomen in one foetus of the 6.3 mg/kg group. In one foetus of the 20.0 mg/kg group both renal pelves were dilated.
Other effects:
no effects observed
Description (incidence and severity):
The placentae of the live foetuses in all groups were normal in weight and showed no macroscopic abnormalities. The placentae of the dead foetuses were in most cases smaller than those of the live foetuses, and were also anaemic.
Details on embryotoxic / teratogenic effects:
Morphological examination of the offspring in all three treatment groups revealed no abnormal findings, apart from an encephalocele in one foetus of the 2.0 mg/kg group. This was certainly of spontaneous origin, since it was an isolated instance, and did not occur in the foetuses of the medium and high dose groups (6.3 and 20.0 mg/kg). All other morphological findings in both dams and foetuses were within the limits of the spontaneous rate. Only a few animals were affected, and they were distributed at random over the various groups.

Effect levels (fetuses)

Dose descriptor:
NOAEL
Effect level:
6.3 mg/kg bw/day
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
other: dead fetuses, resorption and abortion at 20 mg/kg bw/d
Remarks on result:
other: developmental toxicity

Fetal abnormalities

Abnormalities:
no effects observed

Overall developmental toxicity

Developmental effects observed:
yes
Lowest effective dose / conc.:
20 mg/kg bw/day
Treatment related:
yes
Relation to maternal toxicity:
developmental effects occurring together with maternal toxicity effects, but not as a secondary non-specific consequence of maternal toxicity effects

Applicant's summary and conclusion