Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information

Based on the results from teratogenicity studies in rats (Troy Chemical Corporation BV, 1989) and rabbits (BASF SE, 2016) and from the thorough microscopic analysis of sex organs within the scope of a subchronic repeated dose toxicity study in rats there is no indication that adverse effects on reproduction are likely.

Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

There are no specific experimental studies available concerning fertility. However, there is sufficient information from a teratogenicity study and a subchronic repeated dose toxicity study to conclude that the test substance is very unlikely to impair reproductive capacity:

There were no adverse effects on fetal growth or development in a rat and rabbit teratogenicity study with the test substance (Troy Chemical Corporation BV, 1989; BASF SE, 2016). Furthermore, within the scope of a comprehensive subchronic investigation in rats, microscopic inspection of sex organs (oviducts, uterus, vagina, prostate gland, seminal vesicles, left testis, left epididymis, both ovaries) and the mammary tissue from female animals gave no indication to morphological abnormalities (BASF AG, 2002). Subchronic administration of the test substance was not associated with any behavioral changes that would indicate a lack of a desire to mate. Taken together, the available data do not give any indications that adverse effects on reproduction are likely.

Effects on developmental toxicity

Description of key information

In a teratogenicity study in rats (Troy Chemical Corporation BV, 1989) the NOAEL for maternal toxicity was established at 500 mg/kg bw/day, the NOAEL for developmental toxicity was determined to be 750 mg/kg bw/day, the highest dose tested. There were no treatment-related effects on the pregnancy parameters, and embryos showed no developmental effects even at maternally toxic doses.

In a teratogenicity study in rabbits (MELA Triazine REACH Consortium, 2016), the NOAEL for maternal toxicity was set at 60 mg/kg bw/d. The NOAEL for developmental toxicity was established at 60 mg/kg bw/d. There were no treatment-related effects on the pregnancy parameters, and embryos showed no developmental effects even at maternally toxic doses.

Link to relevant study records

Referenceopen allclose all

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2016-05-04 to 2016-08-05
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: BASF SE, 1729 MCA0
- Expiration date of the lot/batch: 07 December 2016

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: At room temperature (between +15 and +25 °C)
- Stability of the test substance in the solvent/vehicle: 96 hours at concentrations of 0.2 to 50 mg/mL
Species:
rabbit
Strain:
New Zealand White
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories France, 01400 Chatillon sur Chalaronne, France
- Age at study initiation: 17 to 19 weeks old
- Weight at study initiation: 3 to 4.5 kg
- Housing: Females were individually housed in composite plastic and metal cages in compliance with European Regulations (Directive 2010/63/EU)
- Diet: ad libitum, Pelleted complete rabbit diet (3409 CRL, KLIBA) sterilised by irradiation and analysed for chemical and bacterial contaminants.
- Water: ad libitum, Softened and filtered (0.2 µm) mains drinking water ad libitum (via an automatic watering system or via bottles).
- Acclimation period: 6 days between animal arrival and the start of treatment

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 ± 3
- Humidity (%): > 35 %
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The test item was prepared as a solution in the vehicle at concentrations of 2, 6 and 18 mg/mL according to Standard Operating Procedures of the Test Facility. It was prepared at least twice weekly. For each concentration, the test item and then the vehicle were weighed and then stirred magnetically in a container until the test item was completely dissolved.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Duplicate samples were taken (2 x 2 mL) according to the above table from each formulation, on a single occasion during the treatment of the main study phase only (except for the group 2 formulation which was taken on additional occasions for accuracy (middle) only due to an initial achieved concentration not in agreement with the theoretical values. The samples were stored at room temperature (between +15 and +25 °C). One set of samples (1 x 2 mL) was analysed at the Test Facility using a validated method.
The duplicate formulation sample (1 x 2 mL) for the nominal 2 mg/mL concentration taken from the first preparation was analysed due to an out of specification value for the first aliquot. Remaining samples were discarded at the end of the stability period.
All analyzed samples for formulations prepared at nominal concentrations of 2, 6 and 18 mg/mL of 2,2’,2”-(hexahydro-1,3,5-triazine-1,3,5-triyl)triethanol in vehicle water for injection, taken from each preparation on the first day of treatment, including the vehicle, were in agreement with acceptance criteria (± 10 %) except formulation of the low dose group at 2 mg/mL. The deviations from the nominal concentrations of Mid and High dose group ranged between -4.2 % and 7.8 % and were thus within acceptance criteria.
No test item was detected in the vehicle sample.
The first formulation for group 2 (Low dose) was outside acceptance criteria as confirmed by duplicate samples analysis. New preparation was made on first day of treatment which was outside acceptance criteria with a deviation of -13.6 %. On fourth day of treatment the preparation method was improved by addition of vehicle by portion in a beaker and stirred manually between each addition. This last preparation of the formulation was slight below the acceptance criteria -10.5 % which was considered acceptable.
Details on mating procedure:
- Impregnation procedure: purchased timed pregnant
Duration of treatment / exposure:
From day 6 to day 28 of gestation
Frequency of treatment:
once daily
Duration of test:
Until termination on day 29
Dose / conc.:
20 mg/kg bw/day (actual dose received)
Dose / conc.:
60 mg/kg bw/day (actual dose received)
Dose / conc.:
180 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
22
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Doses were selected based on the results of the DRF phase.
Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: at least twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: Days 0, 3, 6, 9, 12, 15, 18, 21, 24, 27 and 29 of gestation

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 29
- Organs examined: ovaries and uterus; abnormal organs detected via macroscopic examination
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
Fetal examinations:
- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: all live foetuses per litter
- Skeletal examinations: Yes: all live foetuses per litter
- Head examinations: Yes: half per litter
Statistics:
Statistical analyses were performed by the Provantis data acquisition system, where appropriate, as follows:
The best transformation for the data (none, log or rank) was determined depending upon
- the normality of the data distribution tested by the Shapiro-Wilk's test
- the homogeneity of the variances across groups tested by the Bartlett's test.
Non- or log-transformed data were analysed by parametric methods.
Rank transformed data were analysed using non-parametric methods.
Data were then analysed to test for a dose-related trend to detect the lowest dose at which there is a significant effect, based on the Williams test for parametric data or the Shirley's test for non-parametric data.
Homogeneity of means was assessed by analysis of variances (ANOVA) for parametric data or Kruskal-Wallis test for non-parametric data.
If no trend was found and means are not homogeneous, the data were analysed by parametric or non-parametric Dunnett's test to look for significant differences from the control group.
The number of resorptions, number of dead foetuses and all litter-based percentages were analysed using non-parametric methods, i.e. Kruskal-Wallis test followed by non-parametric Dunnett’s test if the Kruskal-Wallis is significant.
Selected incidence data were analysed using a chi2 test for all groups followed by Fisher’s two-tailed test with Bonferroni correction for each treated group versus the control if the chi2 is significant.
Microsoft Excel ® (2003 or higher) was employed to present certain results.
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Consistent with an effect on food consumption, there was a test-item related higher frequency of reduced/no faecal output in 180 mg/kg/day treated group compared with other groups from G 6 to G 29.
There was no other treatment related clinical signs in any group.
Incidental clinical signs such as sore, scab, pale faeces and red vaginal discharge were observed in the groups given the vehicle, 60 and/or 180 mg/kg/day.
Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Description (incidence):
There was no treatment-related mortality in any group.
Female no. 111 from the control group was sacrificed on G 16 for ethical reason after trauma that occurred during the dosing procedure.
One female given 20 mg/kg/day (no. 135) was found dead on G 22 after treatment. Just after treatment, red traces were noted on the dosing tube and the female had laboured breathing. At necropsy, all right lobes of the lungs were uncollapsed and had many dark foci. Transparent fluid was also present in the thoracic cavity. Therefore the cause of death was considered to be associated with the administration procedure (gavage error).
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
There was a treatment-related and statistically significant lower overall mean body weight gain for females given 180 mg/kg/day between G 6 and G 29 when compared with the control. The effect was principally due to a body weight loss during the first 3 dosing days (-177.4 g between G 6 and G 9). Recovery was noted from G 9 but absolute mean body weight tended to remain slightly lower (approximately 5 %) through to termination.
There was no obvious test item-related effect on body weight gain in the other groups.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
There was an overall dose-related and statistically significant lower mean food consumption for females given 60 and 180 mg/kg/day between G 6 to G 29 (130.1 and 93.3 g/day, respectively) when compared with control group (145.3 g). The effect on food consumption at 180 mg/kg/day was more pronounced from G 6 to G 15.
There was no obvious treatment related effect on mean food consumption at 20 mg/kg/day.
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:
effects observed, non-treatment-related
Description (incidence and severity):
Consistent with lower mean foetal weight, mean gravid uterus weight was slightly (not statistically significant) lower in the 180 mg/kg/day group compared with the control.
There was no test item-related effect on mean gravid uterus weight in the lower dose groups.
Gross pathological findings:
no effects observed
Description (incidence and severity):
There were no treatment-related maternal macroscopic changes.
There were 6, 9, 8 and 5 females given vehicle, 20, 60 and 180 mg/kg/day with one or several cysts on the oviduct(s). These findings are part of the background of changes and incidental.
Other incidental findings were observed such as abnormal shaped uterus, sore/crust and irregular surface of the liver.
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
Description (incidence and severity):
The pre-implantation data (corpora lutea count, number of implantations and the corresponding percentage pre-implantation loss) were comparable in all groups. There was no test item-related effect on embryo-foetal survival in any group.
Total litter losses by resorption:
no effects observed
Early or late resorptions:
not examined
Dead fetuses:
no effects observed
Changes in pregnancy duration:
no effects observed
Description (incidence and severity):
Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): no effects observed
Changes in number of pregnant:
no effects observed
Description (incidence and severity):
There were 19, 21, 21 and 19 pregnant females at terminal caesarean in the control, 20, 60 and 180 mg/kg/day groups, respectively, all of which had viable foetuses.
Key result
Dose descriptor:
NOAEL
Effect level:
60 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
clinical signs
body weight and weight gain
Fetal body weight changes:
effects observed, treatment-related
Description (incidence and severity):
Mean foetal weight was slightly but statistically significantly lower in the 180 mg/kg/day group (38.3 g) when compared with the control (42.2) and below the historical control data range of main studies (39 g – 42.1 g). Dam No. 184, which lost 96 g from G 6 to G 29, had ten considerably light foetuses with multiple malformations.
There was no test item-related effect on mean foetal weight in the lower dose groups.
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): not examined
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Description (incidence and severity):
There was no test item-related influence on foetal sex ratio in any group.
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):
There were 2 (2), 1(1) and 1(1) foetuses (litters) with external malformation in the 180, 60 and 20 mg/kg/day groups, respectively, compared with none in the control group, none of which were attributed to the test item.
In the 180 mg/kg/day group, one foetus (dam no. 168) had an omphalocele and the other (dam no. 188) had a malrotated hindlimb. The foetus in the 60 mg/kg/day group (dam no. 148) had spina bifida and the other in the 20 mg/kg/day group (dam no. 144) had anencephaly.
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
There were 1 (1), 3 (3) and 11 (2) foetuses (litters) with skeletal malformations in the control, 60 and 180 mg/kg/day groups, compared with none in the 20 mg/kg/day group, none of which were attributed to the test item.
The foetus in the control group (dam no. 106) and one from the 60 mg/kg/day group (dam no. 159 had fused sternebrae. The second foetus from the 60 mg/kg/day group (dam no. 148) had confirmation of the spina bifida noted externally in the lumbar/sacral area and the other (dam no. 152) had multiple abnormalities of the cervical and thoracic vertebrae leading to scoliosis. In the 180 mg/kg/day group, 10 of the malformed foetuses came from a single litter (dam no. 184) all of which had a syndrome of atypical incomplete ossification affecting one or more regions of the skeleton (skull, sternebrae, ribs, scapulae, forelimbs and hindlimbs bones and cervical, sacral, lumbar or caudal vertebrae). The other foetus in the 180 mg/kg/day group (dam no. 177) had multiple abnormalities of the cervical vertebrae.
There was a higher incidence of delayed ossification of the extremities in the 180 mg/kg/day group when compared with other groups such as incomplete or unossified forepaw bones (metacarpal and phalanx), hindpaw (metatarsal, phalanx and tarsal bone) and stennebrae.
The incidences of the remaining skeletal anomalies and variations were comparable with the concurrent control and/or historical control data and were considered incidental.
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
There were 3 (3) and 1(1) foetuses (litters) with visceral malformations in the 20 and 180 mg/kg/day groups, respectively, compared with none in the control and 60 mg/kg/day groups, none of which were attributed to the test item.
In the 180 mg/kg/day group, the foetus (dam no. 184) had marked dilated renal pelvis. In the 20 mg/kg/day group, one foetus (dam no. 139) had a three-chambered heart with dilated aortic arch and atretic pulmonary trunk, the second (dam no. 138) had a unilateral retinal fold and the other (dam no. 144) had gross disruption of the brain associated with the anencephaly noted externally.
Other less severe visceral anomalies occurred without dose-dependency or are part of the background of changes noted for this strain of rabbit.
Details on embryotoxic / teratogenic effects:
Foetal examinations revealed a total of 1 (1), 3 (3), 3 (3) and 13 (4) malformed foetuses (litters) in the control, 20, 60 and 180 mg/kg/day groups, respectively, none of which were attributed to the test item due to the lack of any pattern or dose-related increased incidence of any of the findings, which are for the majority also part of the historical control data. In addition, there was no other evidence of a teratogenic potential of the test item with respect to embryo-foetal viability.
Key result
Dose descriptor:
NOAEL
Effect level:
60 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
fetal/pup body weight changes
Key result
Developmental effects observed:
no
Relation to maternal toxicity:
developmental effects as a secondary non-specific consequence of maternal toxicity effects

The numbers of foetuses (litters) submitted to the different examinations were as follows:

Group

1

2

3

4

External examination

178 (19)

209 (21)

200 (21)

181 (19)

Internal examination

 

 

 

 

body

178 (19)

209 (21)

200 (21)

181 (19)

head

83 (19)

97 (20)

96 (21)

86 (19)

Skeletal examination

 

 

 

 

body

178 (19)

209 (21)

200 (21)

181 (19)

head

95 (19)

112 (21)

104 (21)

95 (19)

 

Summary of malformations – Individual descriptions:

Dose level (mg/kg/day)

Female number

Foetus number(s)

Malformation(s)#

0

106

9

Sternebra 3rdto 5thfused

20

138

3

Retinal fold, right eye

139

5

Heart, Three-chambered: presence of a single ventricle and two atria, and right A-V valve appeared closed.

Dilated aortic arch with atretic pulmonary trunk and ductus arteriosus

144

4

Anencephaly with malformed brain: gross disruption with left and right hemispheres misshapen and lying above midbrain

60

148

9

Spina bifida with 4thto 6thlumbar arches and 1stto 3rdsacral arches malpositioned

152

5

Vertebra, multiple abnormalities: 1stcervical centrum bipartite, 4thand 5thcervical centra fused and misshapen, 6thand 7thcervical centra small and hemicentric; odontoïd process unossified; 1stthoracic centrum small and fused with 2ndthoracic centrum, 2ndthoracic centrum misaligned; 1stbilateral cervical arches misshapen and right incomplete ossification, 5thand 7thleft cervical arches small, 5thright cervical arch absent, 2ndleft thoracic arch small; scoliosis

159

4

Sternebra 2ndto 5thfused

180

168

10

Omphalocele

177

9

Cervical vertebra, multiple abnormalities: 2ndcentrum small, hemicentric and fused with odontoïd process; 2ndbilateral arches small

184

2

Skull, Multiple abnormalities: all bones atypical incomplete ossification and misshapen

Vertebral column, multiple abnormalities: from 3rdcervical to 14thcaudal, all sternebrae and scapulae atypical incomplete ossification

3

Multiple skeletal abnormalities: scapulae, forelimb bones, hindlimb bones, vertebra column from 2ndcervical to 14thcaudal,  ribs and sternebrae atypical incomplete ossification

4

Vertebra, multiple abnormalities: 6thand 7thlumbar and 1stsacral atypical incomplete ossification

5

Vertebra, multiple abnormalities: 3rdcervical to 14thcaudal atypical incomplete ossification

Kidney, Dilated renal pelvis (right), marked

6

Vertebral column, multiple abnormalities: from 2ndcervical to 14thcaudal and all sternebrae atypical incomplete ossification

7

Vertebral column, multiple abnormalities: from 2ndcervical to 14thcaudal and all sternebrae atypical incomplete ossification

8

Vertebra, multiple abnormalities: 2ndcervical to 3rdcaudal atypical incomplete ossification

9

Vertebra, multiple abnormalities: 3rdcervical to 14thcaudal atypical incomplete ossification

10

Skull, multiple abnormalities: zygomatic arches, frontals and parietals atypical incomplete ossification; interparietal unossified

Vertebra, multiple abnormalities: 3rdcervical to 14thcaudal atypical incomplete ossification

11

Vertebral column, multiple abnormalities: from 3rdcervical to 3rdcaudal and scapulae atypical incomplete ossification

188

14

Malrotation of right hindlimb

#: including external, visceral and skeletal examinations.

 

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
EU Method B.31 (Prenatal Developmental Toxicity Study)
Deviations:
yes
Remarks:
Rats were only dosed from Day 6 to 15 rather than 6 to 19 of gestation as required by current guidelines.
Qualifier:
according to
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
Deviations:
yes
Remarks:
Rats were only dosed from Day 6 to 15 rather than 6 to 19 of gestation as required by current guidelines.
GLP compliance:
yes
Limit test:
no
Specific details on test material used for the study:
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Stability under test conditions: stable in distilled water over a 29-day period
Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Iffa Credo, Belgium
- Weight at study initiation: Group mean bodyweights for females at the start of the study: 228 to 309 grams
Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
- Duration of exposure: 10 consecutive administrations from day 6 to day 15 of pregnancy
- Postexposure period : The rats were untreated from Day 16 to terminaton on Day 20
- Vehicle: Distilled water
- Total volume applied: 10 mL/kg bw/day, volume adjusted daily based on individual body weight
- Controls: Vehicle – distilled water
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Dose solutions were prepared twice during the study, and content of test article in all dose solutions was verified by analysis.
Details on mating procedure:
Males and females were mated in a ratio of one male to two females. Mating was detected by observation of a sperm-positive vaginal smear. The day of observation of sperm was termed day 0 of pregnancy.
Duration of treatment / exposure:
days 6 through 15 of pregnancy
Frequency of treatment:
once daily
Duration of test:
until terminaton on Day 20
Remarks:
Doses / Concentrations:
250, 500 and 750 mg/kg bw/d
Basis:
actual ingested
No. of animals per sex per dose:
24 females per group, 4 groups
Control animals:
yes, concurrent vehicle
Maternal examinations:
- Body weight: Yes. Animals were weighed on gestation days 0, daily from days 6 to 15, and on day 20
- Food consumption: Yes. Recorded for each animal gestation days 0 to 6, 6 to 11, 11 to 15, and 15 to 20
- Clinical signs: Yes. Recorded daily from gestation day 0 to 20.
- Haematology and spleen weight: No
Ovaries and uterine content:
At necropsy, the following parameters were assessed for each surviving dam:
1 Weight of gravid uterus;
2 Abnormalities of major maternal organs;
3 Number of corpora lutea; and
4 Number and distribution of implantation sites with classification as early resorption, late resorption, dead foetus, or live foetus.

One animal in the 750 mg/kg bw/day was killed prematurely because of poor clinical condition. The uterine content of the decedent animal was examined for items 2, 3, and 4 above.
Fetal examinations:
For each surviving dam the following foetal parameters were determined:
- Live foetal weight;
- Foetal sex; and
- External abnormalities of foetuses.
One half of the living foetuses processed for examination of skeletal shape, size, and extent of ossification.
Remaining foetuses were fixed in Bouin’s fluid and examined by a combined sectioning/dissection technique. Serial sections of the head were examined. The thorax and abdomen were examined by microdissection, including heart, kidneys, and all major organs and blood vessels.
Structural congenital abnormalities were classified as major if they impaired or could potentially impair survival or fitness. Other defects were classified as minor abnormalities. Commonly observed variations from normal ossification were recorded as variants.
Statistics:
Group mean maternal bodyweights, bodyweight gain and food consumption, number of corpora lutea, number of live foetuses, total number of implantation sites, and foetal bodyweights per litter were subjected to analysis of variance and Student’s t test.
Pre-implantation and post-implantation loss, percentages of male and female foetuses per litter, and percentages of foetuses exhibiting abnormalities were analyzed by Kruskal-Wallis test.
Details on maternal toxic effects:
Details on maternal toxic effects:
Mortality:
One female in the 750 mg/kg bw/day group was killed on day 14 of pregnancy because of poor clinical condition. No other premature deaths occurred.
Clinical signs:
The decedent from the 750 mg/kg bw/day exhibited slight emaciation and piloerection, pale extremities, laboured breathing, rales, and red staining around mouth and nose on the day it was put to death. Previously, the only observed clinical abnormalities in this animal had been post-dose salivation.
Post-dose salivation was observed frequently in the 750 mg/kg bw/day group, particularly towards the end of the dosing period. Rales, wheezing, laboured breathing, or tachypnoea were observed occasionally at 500 and 750 mg/kg bw/day at the end of the dosing period.
No treatment-related clinical signs occurred in the 250 mg/kg bw/day group.
Bodyweight:
Mean bodyweight gain in the 750 mg/kg bw/day group was significantly reduced in comparison to the control group from initiation of treatment. Bodyweight gain was not affected by treatment at lower levels.
Bodyweight gain after treatment ceased was similar to the control group.
Food consumption:
Mean food consumption in the 750 mg/kg bw/day group was significantly reduced in comparison to the control group during treatment. In particular, the decedent animal showed a marked reduction in food consumption during dosing.
Food consumption was not affected at other dose levels.
Maternal necropsy:
Stomach lesions, i.e., ulceration and/or scar tissue on mucosa, occurred in 14 of 24 females in the 750 mg/kg bw/day group, including the decedent animal. No abnormalities were found in the 250 or 500 mg/kg bw/day groups.
Pregnancy data:
The number of pregnant females at sacrifice in the 0, 250, 500, and 750 mg/kg bw/day groups was 21, 22, 24, and 22, respectively. No significant treatment-related inter-group differences in numbers of corpora lutea, implantation sites, live foetuses, or pre-and post-implantation loss occurred. Foetal bodyweights, uterine weights, and foetal sex ratios were similar for all groups.
Key result
Dose descriptor:
NOAEL
Effect level:
500 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
body weight and weight gain
food consumption and compound intake
Details on embryotoxic / teratogenic effects:
Details on embryotoxic / teratogenic effects:
Foetal examination:
No treatment-related major abnormalities were observed in foetuses from any group.
Major abnormalities did occur in three foetuses, i.e., one from the control group and one from the 750 mg/kg bw/day group had an umbilical hernia, and one from the 750 mg/kg bw/day exhibited encephaly, microphthalmia, protruding tongue, and cleft palate. These abnormalities occasionally occur spontaneously in Sprague-Dawley derived rats and therefore were not considered to be treatment-related.
There were no statistically significant differences in incidences of minor skeletal abnormalities between treated groups and controls.
Some intergroup differences in the incidences of minor skeletal abnormalities were observed but these did not achieve statistical significance and were considered not to be treatment-related. The overall incidence of foetuses with minor skeletal abnormalities was greater in the treated groups than the controls with the differences following an apparent dose-related trend. These differences were principally the result of increased incidences of retarded ossification of thoracic vertebral centra and the presence of vestigial ribs.
The retarded ossification in thoracic vertebral centra was not part of a consistent trend of developmental retardation because some variants of ossification, such as the presence of centers for ossification of phalanges, showed advanced ossification in the treated groups, and foetal bodyweights were not affected by treatment.
The dose-related increase in vestigial ribs was also not considered to be treatment related, because the incidence of this abnormality was only slightly greater than historical background incidence in Sprague-Dawley OFA-SD (IOPS-Caw) rats.
Key result
Dose descriptor:
NOAEL
Effect level:
>= 750 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Key result
Developmental effects observed:
no

Table 1: Table of maternal effects:

 

Dose Level, mg/kg bw/day

Parameter

0

250

500

750

Number of dams examined

24

24

24

24

Clinical findings:
Piloerection

Râles

Slight emaciation

Laboured breathing

Red staining on muzzle

Post-dose salivation

Wheezing

Tachypnoea

Alopecia both forelimbs

 

Macroscopic pathology:

Grossly distended stomach with gas

Ulcerated and/or scar tissue on stomach mucosa

Scar tissue on non-glandular stomach mucosa.

 

0

0

0

0

0

0

0

0

0

 

 

0

 

0

 

0

 

0

0

0

0

0

0

0

0

0

 

 

0

 

0

 

0

 

0

2B

0

0

0

0

1C

1

1

 

 

0

 

0

 

0

 

1A

2A

1A

2A

2A

21A

0

0

0

 

 

1A

 

14A

 

1A

 

Mortality of dams

0

0

0

1

Group mean weight (g)

day 0
day 6
day 9
day 12

day 15
day 20

day 20C

 

269

294

309

319

341

409

322

 

272

296

311

326

347

417

324

 

268

293

304

320

340

413

317

 

271

295

301

311

331

397

307

Group mean weight gain (g)

day 6 to 9
day 6 to 12

day 6 to 15
day 6 to 20

day 6 to 20

 

12

25

48

115

28

 

15

30

51

121

28

 

12

28

47

120

24

 

6**

16**

36**

102*

12***

Food consumption [g] (mean/animal/day)H

day 0 to 6
day 6 to 11

day 11 to 15
day 15 to 20

 

 

23.0

25.3

27.7

27.7

 

 

23.0

25.2

27.6

27.7

 

 

23.7

23.9

26.7

27.6

 

 

23.1

18.3***

23.5***

27.0

Pregnancy
Females per group
Total number pregnant
Pregnancy rate as % of mated

 

24

21

87.5

 

24

22

91.7

 

24

24

100

 

24

22

91.7

A Including decedent animal.

B One animal had pre- and post-dose rales

C One animal had pre- and post-dose wheezing

* = significantly different from control, p < 0.05, Student’s t test

** = significantly different from control, p < 0.01, Student’s t test

*** = significantly different from control, p < 0.001, Student’s t test

Table 2: Table of litter response:

 

Dose Level, mg/kg bw/day

Parameter

0

250

500

750

Mean number of corpora lutea

18.3

18.8

18.5

18.2

Mean number of implantations

14.4

15.1

15.8

15.1

Mean number of live foetuses

13.7

14.6

15.2

14.5

Mean Pre-implantation loss (%)

21.3

20.2

14.2

18.1

Mean Post-implantation loss (%)

4.8

2.8

3.6

3.0

Mean Sex ratio (male to female)

54 / 46

50 / 50

49 / 51

53 / 47

Major foetal abnormalities (%)

Encephaly, microphthalmia, protruding tongue, cleft palate

Umbilicus : hernia

0.3

0

 

1.3

0.0

0

 

0

0.0

0

 

0

0.5

0.3

 

0.3

Minor foetal abnormalities (%)

Total minor total skeletal abnormalities only (%)

Vertebrae: thoracic centra, one or more retarded ossification

Rib: uni-or bilateral vestigial 14th

13.8

12.2

2.1

 

8.1

16.3

17.5

2.2

 

11.0

11.6

24.8

4.9

 

13.0

7.6*

24.0

8.5

 

15.2

Mean foetus weight [g]

3.78

3.79

3.78

3.72

Mean uterine weight [g]

87.2

82.9

96.5

89.4

* = significantly different from control, p < 0.05, Student’s t test

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
60 mg/kg bw/day
Species:
rabbit
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

In a prenatal developmental toxicity study, that was performed according to EPA guideline OPPTS 870.3700 (§83-3; Prenatal Developmental Toxicity Study) and EU Method B.31 (Prenatal Developmental Toxicity Study), artificially inseminated female Sprague-Dawley rats were dosed orally by gavage at 0, 250, 500 or 750 mg/kg bw/day from day 6 to day 15 of pregnancy (Troy Chemical Corporation BV, 1989). In this study, maternal toxicity was evident at 750 mg/kg bw/day: mortality (1 female was killed moribund), respiratory disturbance reflected in clinical signs such as rales, reduced bodyweight or reduced weight gains and lower food consumption and stomach lesions were evident at necropsy. However, there were no treatment-related effects on the pregnancy parameters and embryos showed no developmental effects even at maternally toxic doses. The NOAEL for maternal toxicity was 500 mg/kg bw/day. The NOAEL for developmental toxicity was determined to be 750 mg/kg bw/day, the highest dose tested.

Rabbit

A prenatal developmental toxicity study was performed according to OECD guideline 414. The test item was administered by daily oral gavage at dose levels of 20, 60 and 180 mg/kg/day to groups of 22 mated female New Zealand White rabbits from days 6 to 28 of gestation inclusive. A control group received the same dose volume (10 mL/kg) of the vehicle water for injection. Clinical conditions, body weight and food consumption were monitored throughout the study. The females were submitted to a caesarean examination on day 29 of gestation and litter parameters were recorded. At necropsy the females were examined macroscopically and the gravid uterus was weighed. All foetuses were weighed, examined for external and visceral abnormalities and sexed. The head of approximately half of the foetuses in each litter were fixed for internal examination by serial sectioning. The eviscerated carcasses of all foetuses were processed for skeletal examination.

Oral administration to the pregnant New Zealand White rabbit was associated with relatively severe maternal toxicity in the high dose group including a lower overall mean body weight gain and a lower mean food consumption. The intermediate dose of 60 mg/kg/day was a No Observed Adverse Effect Level for maternal toxicity.

Consistent with the degree of maternal toxicity, lower mean foetal weight associated with delayed ossification of several bones was noted in the 180 mg/kg/day group. It is known from the literature that reductions in foetal body weights and delays in ossification frequently occur concurrent with reduced maternal food consumption and maternal body weights, similar to the current study. There was no evidence of a teratogenic potential of 2,2’,2”-(hexahydro-1,3,5-triazine-1,3,5-triyl)triethanol in any group. The No Observed Adverse Effect Level for embryo-foetal development was therefore considered to be 60 mg/kg/day in view of the effects associated with lower foetal weight.

Justification for classification or non-classification

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. No teratogenic effects were observed in rats and rabbits. Furthermore, no evidence is given for reproducitve toxicity in subchronic repeated dose toxicity studies and the two developmental toxicity studies. As a result, the substance is not considered to be classified for reproductive or developmental toxicity under Regulation (EC) No 1272/2008, as amended for the ninth time in Regulation (EU) No 2016/1179.