Ethylenediamine, ethoxylated and propoxylated

Toxicity to reproduction

Administrative data

Endpoint:

screening for reproductive / developmental toxicity

Remarks:

based on test type (migrated information)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: 1a: Guideline study (GLP)

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2007

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)

Deviations:

no

Remarks:

not mentioned

Principles of method if other than guideline:

not applicable

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Test material

Test material information

Constituent 1

Details on test material:

Voranol RA 800; Molecular Mass: (average Mn = 280 g/mol)
Units of Propylene oxide, approx: 70-90%, Units of ethylene oxide: approx 10-30%
Lot #: UB06083011
Colourless viscous Fluid

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

Animals were supplied by "Charles River, Sulzfeld, Germany" and were about 10 (males) and 15 (females) weeks old and had an initial mean wights of 351 and 197 g, respectively.
Housing singly in Makrolon(R) cages Type IIIh on wood granules. All animals or the study were kept in the same room with no other animals from other studies and in an 20 Pa overpressure environment at 23°C, 55 +/-5% relative humidity and a 12 hour light/dark cycle and low-noise music.
Diet was stadard diet "Provimi Kliva 3883.0.15" pellets supplied by Provimi Kliva SA, Kaiseraugst, Switzerland ad libitum. Water was provided ad libitum.

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Details on exposure:

vehicle: demineralized water
administration volume: 10 ml/kg bw

Details on mating procedure:

Pairing was performed overnight by placing one F0 female animal together with one F0 male rat. If sperm was detected in the vaginal smear taken on the morning following mating, this day was regarded as day 0 of gestation. Animals were paired daily during the 2-week mating and one week remating period. Females in which insemination had not been detected by the end of the 2-week mating period, were mated for another week with another male of the respective dose group which had successfully inseminated a female paired with it. F0 females found sperm-positive after the first matings but where body weight gain did not indicate pregnancy by the end of the 2-week mating period were paired again for 7 days during the remating period.

Analytical verification of doses or concentrations:

no

Details on analytical verification of doses or concentrations:

not applicable

Duration of treatment / exposure:

up to 58 days.

Frequency of treatment:

daily

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day

Remarks:

Basis:actual ingested

Doses / concentrations 2

Dose / conc.:

100 mg/kg bw/day

Remarks:

Basis:actual ingested

Doses / concentrations 3

Dose / conc.:

300 mg/kg bw/day

Remarks:

Basis: actual ingested

Doses / concentrations 4

Dose / conc.:

1 000 mg/kg bw/day

Remarks:

Basis:actual ingested

No. of animals per sex per dose:

12 male and 12 female Wistar rats/dose and control group

Control animals:

yes, concurrent vehicle

Details on study design:

ADMINISTRATION/EXPOSURE

- Treatment: The male F0 rats were each given ethylenediamine, ethoxylated and propoxylated (NLP#6) or vehicle (demineralized water) once daily for 2 weeks prior to mating, during the following mating and remating period and up to the day before necropsy (necropsy on day 40 of the study; one male which was sacrificed moribund was necropsied on day 36 of the study). NLP#6 or vehicle was administered to each of the female F0 rats once daily for 2 weeks prior to mating, during the subsequent mating and remating period, during gestation and up to the day before necropsy. Necropsy was either performed on day 4 to 7 p.p. in females which delivered, or in inseminated females which did not deliver some days after the expected day of delivery; i.e. the F0 females were treated individually differently for 42 to 58 days.

Positive control:

No positive control

Examinations

Parental animals: Observations and examinations:

Clinical examination, mortality, body weight and food intake.
Urinalysis: Urine was collected for about 16 hours from 5 rats per dose and sex*. in males this was done on the first 5 males per groups on day 30. In females this was done in those five females, which had been necropsied first (day 45) mostly after pups had been killed.
Animals were fasted during collection period and water was offered for ad libitum consumption. The following parameters were determined:

- Quantitatively: Volume, density, protein per vol., creatine per vol.
- Semiquantitatively: pH, glucose, protein, blood, bilirubin, ketone bodies, urobilinogen
- microscopy of sediment
*These examinations were done as urinalysis in a subacute study (Report No. AT03512) had been erroneously incomplete.

Sperm parameters (parental animals):

- testes
- epididymides
- histopatholocigal evaluation

Litter observations:

- Birth and viability
- L itter siye at birth and the live index
- Sex ratio
- Clinical observation during first days of lactation
- Pup weights
- Gross pathological changes in the F1 pups.

Postmortem examinations (parental animals):

ORGANS EXAMINED AT NECROPSY (MACROSCOPIC AND MICROSCOPIC) F0:
Organ weights: testes and epididymides (right and left organs separated)
Histopathology: testes, epididymes, and ovaries with oviducts in the control and high dose group.
Implantation sites in the uterus were counted at necropsy while number of corpora lutea in the ovaries was recorded during histopathological investigations (only control and high dose group

Postmortem examinations (offspring):

PARAMETERS ASSESSED ON F1:
- clinical signs
- number of live and dead pups
- sex of pups
- body weights
- external macroscopical investigations (including apparent malformations)

Statistics:

statistical significance was tested using the TASC-System; Analysis of variance (ANOVA) and in case of significant results DUNNETT´s test; 2/N CHI-Square test; in case of sigificant differences Fishers´s exact test with Bonferroni correction.

Reproductive indices:

- Insemination index (%): # of Sperm positive females/ # of females co-housed with a male
- Fertility index (%): # of pregnant females / # of sperm positive females
- Gestation index (%): # of females wiht live pups / # of pregnant females

Offspring viability indices:

- Live birth index (%): # of live pups at birth / total # of pups born
- Viability index (%): # of live pups on day 4 / # of live pups born

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

no effects observed

Dermal irritation (if dermal study):

not specified

Mortality:

no mortality observed

Description (incidence):

During the study no animal died in the study groups up to 1000 mg/kg.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

There were no decreases in body weights or body weight gain up to and including 1000 mg/kg in males and females during the premating period. Lactating 1000 mg/kg females exhibited a mild (6-7%) body weight depression (p<0.05), while body weight gain during lactation was unaffected.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Food efficiency:

not examined

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

effects observed, non-treatment-related

Description (incidence and severity):

The semiquantitative urinalyses including the sediment showed no toxicologically relevant findings up to and including 1000 mg/kg

Behaviour (functional findings):

no effects observed

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

not examined

Other effects:

not examined

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:

not specified

Reproductive function: sperm measures:

no effects observed

Reproductive performance:

no effects observed

Details on results (P0)

Mild body weight depression during lactation at 1000 mg/kg were reported. The registrant considers the mild body weight depression of 6 to 7% reported in the 1000 mg/kg bw/day in female during lactation as not adverse as the body weight gain during lactation was unaffected. An adverse effect is usually considered to be equal to or greater than 10% reduction in body weight.

Effect levels (P0)

1

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Sex:

female

Basis for effect level:

other: see 'Remark'

2

Dose descriptor:

NOAEL

Effect level:

300 mg/kg bw/day (nominal)

Sex:

female

Basis for effect level:

other: NOAEL as reported in the study report due to body weight depression during lactation at 1000 mg/kg. However, see comment from registrant.

3

Dose descriptor:

NOEL

Effect level:

300 mg/kg bw/day (nominal)

Sex:

female

Basis for effect level:

other: Salivation at 1000 mg/kg bw/day

4

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Sex:

male

Basis for effect level:

other: limit dose

5

Dose descriptor:

NOEL

Effect level:

300 mg/kg bw/day (nominal)

Sex:

male

Basis for effect level:

other: Salivation at 1000 mg/kg bw/day

Results: P1 (second parental generation)

Target system / organ toxicity (P1)

Critical effects observed:

not specified

Results: F1 generation

General toxicity (F1)

Clinical signs:

no effects observed

Mortality / viability:

no mortality observed

Body weight and weight changes:

no effects observed

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings:

no effects observed

Effect levels (F1)

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

1 000 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: reproduction/developmental toxicity

Target system / organ toxicity (F1)

Critical effects observed:

not specified

Results: F2 generation

Target system / organ toxicity (F2)

Critical effects observed:

not specified

Overall reproductive toxicity

Reproductive effects observed:

not specified

Any other information on results incl. tables

Survival rate of F0 animals was not reduced by treatment with ethylenediamine, ethoxylated and propoxylated at a dose level up to and including 1000 mg/kg bw/day.

Salivation directly after the administration was observed in both genders at the 1000 mg/kg dose level but an adverse effect is not concluded from this finding.

The lowest-observed-effect-level (LOEL) and the no-observed-effect level (NOEL) for salivation after administration were as follows:

Males: LOEL: 1000 mg/kg bw/day

NOEL: 300 mg/kg bw/day

Females LOEL: 1000 mg/kg bw/day

NOEL: 300 mg/kg bw/day

Body weight development was not affected to a toxicologically relevant extent in males at a dose level up to and including 1000 mg/kg. Females of the 1000 mg/kg group revealed a mild body weight gain depression during lactation.

There was no difference in food intake among groups up to and including 1000 mg/kg bw/day.

Urine parameters were not changed up to and including 1000 mg/kg bw/day.

PARAMETERS OF REPRODUCTION

Treatment related effects on reproductive parameters, i.e. insemination, fertility and gestation index, time to insemination, duration of gestation, course of birth, lactation behavior, number of corpora lutea, implantation sites, prenatal loss, litter size, sex ratio of pups, pup mortality (live birth index, viability index), pup body weight, and clinical findings of pups were not evident at a dose level up to and including 1000 mg/kg.

An indication for a teratogenic potential of ethylendiamine, ethoxylated and propoxylated, was not evident in this study at a dose level up to and including 1000 mg/kg bw/day.

PATHOLOGY

Necropsy, evaluation of male reproductive organ weight (testes, epididymides) and histopathology of reproductive organs (testes, epididymides, ovaries and oviducts) revealed no treatment related findings up to and including 1000 mg/kg.

Applicant's summary and conclusion

Conclusions:

The registrant considers the mild body weight depression of 6 to 7% reported in the 1000 mg/kg bw/day in female during lactation as not adverse as the body weight gain during lactation was unaffected. An adverse effect is usually considered to be equal to or greater than 10% reduction in body weight.

Executive summary:

NLP#6 (Ethylenediamine, Ethoxylated and Propoxylated Polyol, molecular mass 280 g/mol) was administered daily via gavage in demineralized water to 12 male and 12 female Wistar rats per dose group, in doses of 0, 100, 300 or 1000 mg/kg body weight for a period of 2 weeks prior to a 2 week mating period. Males were treated then further up to necropsy (4 weeks at a minimum) and females during gestation and lactation up to their necropsy after day 4 p.p. of their pups. Investigations were performed on general tolerance of the test compound by the parental animals as well as on effects on reproduction and early postnatal development of Fl pups. The animals were regularly observed and weighed, food intake as well as urine and reproduction parameters were determined. Selected organs were weighed and organs were subjected to macroscopical and histopathological investigations. The test substance was stable in the vehicle for the duration of use. Formulations given to the rats were prepared appropriately. Survival rate of FO rats was not reduced up to and including 1000 mg/kg. At 1000 mg/kg FO rats exhibited salivation directly after the administration. An adverse effect is not concluded from this finding. In lactating females a mild body weight depression was seen at 1000 mg/kg. There was no difference in food intake among groups up to and including 1000 mg/kg. Urine parameters were not changed up to and including 1000 mg/kg. No adverse effects were seen in the weights of the testes, epididymides or ovaries. Histopathology revealed no evidence of test substance induced changes in male and female rats with respect to testes, epididymides, ovaries and oviducts at 1000 mg/kg. Up to and including 1000 mg/kg no effects on mating behavior, fertility and live birth indices, gestation parameters, number of implantation sites or prenatal loss, litter size, number of pups born, sex ratio, viability of pups and pup weights were seen. There were also no remarkable clinical or necropsy findings in pups and no adverse effect on the course of birth or lactation behavior of the dams in these groups.

Thus, the following no-observed-(adverse) effect levels were determined:

Females (body weight depression during lactation) NOAELand NOEL 300 mg/kg

Males NOEL: 1000 mg/kg NOAEL 300 mg/kg

Reproduction/Developmental Toxicity: NOAEL 1000 mg/kg