Yellow PE 3260

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Link to relevant study records

Reference

Endpoint:

screening for reproductive / developmental toxicity

Remarks:

based on test type

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2011

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)

Version / remarks:

1995

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

other: Han Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan Laboratories, B.V., Kreuzelweg 53, 5961 NM Horst / Netherlands
- Females (if applicable) nulliparous and non-pregnant: [yes/no]
- Age at study initiation: (P) 11 wks; (F1) x wks
- Weight at study initiation: (P) Males: 289-338 g; Females: 182-211 g; (F1) Males: x-x g; Females: x-x g
- Fasting period before study: not specified
- Housing: Individually in Makrolon type-3 cages. During the prepairing period, cages with males were interspersed amongst those holding females to promote the development of regular estrus cycles
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: at least 7 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30-70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12 / 12 fluorescent light
IN-LIFE DATES: From: 09/10/2010 To: 11/11/2010

Route of administration:

oral: gavage

Vehicle:

water

Details on exposure:

First Test Item Administration: Males and Females: Day 1 of pre-pairing

Details on mating procedure:

- M/F ratio per cage:
- Length of cohabitation:
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility.
- Further matings after two unsuccessful attempts: [no / yes (explain)]
- After successful mating each pregnant female was caged (how):
- Any other deviations from standard protocol:

Analytical verification of doses or concentrations:

yes

Duration of treatment / exposure:

Males were treated over a 14-day pre-pairing period and during the pairing period up to
one day before necropsy (ca. 4 weeks). Females were treated throughout the pre-pairing, pairing, gestation and lactation period up to the day 3 post partum (7 weeks).

Frequency of treatment:

Once daily: A standard dose volume of 10 mL/kg body weight with a daily adjustment to the actual body weight was used.

Details on study schedule:

- F1 parental animals not mated until [...] weeks after selected from the F1 litters.
- Selection of parents from F1 generation when pups were [...] days of age.
- Age at mating of the mated animals in the study: [...] weeks

Dose / conc.:

1 000 mg/kg bw/day (nominal)

Dose / conc.:

300 mg/kg bw/day (nominal)

Dose / conc.:

100 mg/kg bw/day (nominal)

Dose / conc.:

0 mg/kg bw/day (nominal)

Remarks:

control group (vehicle, Milli Q-Water)

No. of animals per sex per dose:

Number of animals: 40 males: 10 per group; 40 females: 10 per group

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: The dose levels were selected based on a previous Subacute 28-Day Oral Toxicity (Gavage) Study with Yellow PE 3260/SF in Han Wistar rats, RCC Project Number 632182, using dose levels of 0, 50, 200 and 1000 mg/kg/day, where no adverse effects were detected up to and including the dose level of 1000 mg/kg bw/day.
- Rationale for animal assignment (if not random): Computer-generated random algorithm. In addition body weights (recorded on the day of allocation) were taken into consideration in order to ensure similar mean body weights in all groups.

Parental animals: Observations and examinations:

VIABILITY/MORTALITY: Yes, twice daily.

CLINICAL OBSERVATIONS: Yes
Daily cage-side clinical observations (once daily, during acclimatization and up to day of necropsy). Additionally females were observed for signs of difficult or prolonged parturition, and behavioral abnormalities in nesting and nursing.

BODY WEIGHT: Yes
- Time schedule for examinations: daily from treatment start to day of necropsy.

FOOD CONSUMPTION AND COMPOUND INTAKE:
Males: Weekly during pre-pairing and after pairing periods.
Females: Pre-pairing period days 1 - 8 and 8 - 14, gestation period days 0 - 7, 7 - 14 and 14 - 21 post coitum and lactation period days 1 - 4 post partum.

Litter observations:

The litters were examined for litter size, live births, still births and any gross anomalies. The sex ratio of the pups was recorded. Pups were weighed individually (without identification) on day 1 and 4 post partum.

Postmortem examinations (parental animals):

SACRIFICE
Males were sacrificed after they had been treated for at least 28 days.
If birth did not occur on the expected date (day 21 post coitum), the dam was sacrificed on day 25 post coitum.

GROSS NECROPSY
All parent animals and pups were examined macroscopically for any structural changes. For the parent animals, special attention was directed at the organs of the reproductive system. The number of implantation sites and corpora lutea were recorded for all dams with litters. The uteri of non-pregnant females were placed in a solution of ammonium sulfide to visualize possible hemorrhagic areas of implantation sites

HISTOPATHOLOGY / ORGAN WEIGHTS
Slides of all organs and tissues collected at terminal sacrifice from the animals of the control and high-dose groups were examined by the study pathologist. The same applied to all occurring gross lesions.
Special emphasis was made on the stages of spermatogenesis and histopathology of interstitial cell structure.
Histological examination of ovaries was carried out on any females that did not give birth. In addition, microscopic examination of the reproductive organs of all infertile males was made.
At the scheduled sacrifice, the testes and epididymides of all parental males were weighed separately.

Postmortem examinations (offspring):

SACRIFICE
Dams and pups were sacrificed on day 4 post partum.

GROSS NECROPSY
Dead pups, were examined macroscopically. All parent animals and pups were examined macroscopically for any structural changes

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

All animals survived until the scheduled necropsy.
No adverse effects were noted in males or females at any dose levels.
Treatment with the test item caused orange discoloration of feces in all animals at the dose levels
of 100, 300 and 1000 mg/kg bw/day. This effect was a result of staining ability of the test item
and considered not to be adverse to the animals.

Mortality:

no mortality observed

Description (incidence):

All animals survived until the scheduled necropsy.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

No test item-related effects on body weights or body weight gain of males or females were observed at any dose level.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

No test item-related effects on food consumption of males or females were observed at any dose
level.

Organ weight findings including organ / body weight ratios:

no effects observed

Reproductive performance:

no effects observed

Description (incidence and severity):

No test item-related effects on the relevant reproductive data (mating performance, fertility,
corpora lutea count, implantation rate and post-implantation loss, duration of gestation, litter size
at first litter check or postnatal loss) were observed at any dose level.

The reproduction parameters investigated did not give any indication of any test item-related effects. Mating performance, fertility and duration of gestation were not affected by the treatment with the test item. Relevant reproduction parameters such as mean number of corpora lutea, mean number of implantations per dam, post-implantation losses and litter size were also not affected by the treatment with the test item.
Gross pathology findings were considered to be within the range of normal background lesions. Only findings common in rats of this strain and age have been noted during gross pathology. All of them were within the range of historical control and are reflecting the usual individual variability. No significant findings for organ weights.

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Remarks on result:

other: NOAEL for general toxicity (highest dose level used in the study)

Dose descriptor:

NOEL

Effect level:

1 000 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Remarks on result:

other: NOEL for reproduction/developmental toxicity (the highest dose level used in the study)

Clinical signs:

no effects observed

Description (incidence and severity):

No findings were noted in pups at first litter check or during the first 4 days post partum.
Pups sex ratio was not affected by the treatment with the test item at any dose level.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

No effects were observed on pup body weight and body weight gain during the first four days
post partum at any dose level.

Gross pathological findings:

no effects observed

Description (incidence and severity):

No findings were noted during macroscopic examination of pups at any dose level.

Viabilitiy index was 100% in the control group and at the dose levels 300 and 1000 mg/kg bw/day and 94.5% at the dose level of 100 mg/kg bw/day. This difference in viability index in the 100 mg/kg bw/day was due to five pups which were missing on day 2 post partum. All these pups were from one litter. No further posnatal loss was noted at any dose level. No findings were noted in pups at first litter check or during the first 4 days post partum. Pups sex ratio was not affected by the treatment with the test item at any dose level. No effects were observed on pup body weight and body weight gain during the first four days post partum at any dose level. No findings were noted during macroscopic examination of pups at any dose level.

Dose descriptor:

NOEL

Generation:

F1

Effect level:

1 000 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Remarks on result:

other: NOEL for reproduction / developmental toxicity

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

1 000 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Remarks on result:

other: NOAEL for general toxicity

Reproductive effects observed:

not specified

The only effect seen under the conditions of this test was the discoloration of feces occurring in all animals which received the test item. The NOAEL (No Observed Adverse Effect Level) for general toxicity was established at 1000 mg/kg/bw/day (the highest dose level used in the study).

The NOEL (No Observed Effect Level) for reproduction/developmental toxicity was established at 1000 mg/kg bw/day (the highest dose level used in the study).

Conclusions:

Based on the discoloration of feces occurring in all animals which received the test item, the
NOAEL (No Observed Adverse Effect Level) for general toxicity was established at 1000 mg/kg
bw/day (the highest dose level used in the study).
The NOEL (No Observed Effect Level) for reproduction/developmental toxicity was established
at 1000 mg/kg bw/day (the highest dose level used in the study).

Executive summary:

This study is a valid investigation of the toxicological effects resulting from repeated oral-gavage administration of the test item Yellow PE 3260/SF to rats. Yellow PE 3260/SF was administered in Milli-Q-Water as vehicle at dosages of 100, 300, and 1000 mg/kg body weight/day, and controls received the vehicle only. Yellow PE 3260/SF was administered to male rats for at least 28 days and to female rats for 14 days prior to pairing, through the pairing and gestation periods until the F1 pups reached day 4 post partum.
All animals survived the scheduled study period.
With the exception for discoloration of feces, no other test item-related findings were noted in males or females at any dose level. The orange discoloration of feces was observed in all animals treated with the test item. This finding was a result of staining ability of the test item and was considered not to be adverse.
No indices of a general toxicity were observed during the entire study at any dose level. Food consumption, body weights and body weight gain were not affected by the treatment at any dose level. No macroscopical or histopathological findings, which were related to the treatment, were found in males or females in any group as well no changes of organ weights were observed.
The reproduction and developmental parameters investigated within this study did not give any indication of any test item-related effect.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

Kimisch 1 rated experimental study

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

A valid OECD 421 reproduction/development toxicity screening study has been conducted with the registered substance. The substance proved to be not reprotoxic up to a dose level of 1000 mg/kg bw/d.

The reproduction parameters investigated did not give any indication of any test item-related effects. Mating performance, fertility and duration of gestation were not affected by the treatment with the test item. Relevant reproduction parameters such as mean number of corpora lutea, mean number of implantations per dam, post-implantation losses and litter size were also not affected by the treatment with the test item.

Additional information with regard to this endpoint is available from a vlaid OECD 407 oral subacute toxicity study, in which the following fertility related endpoints were monitored:

- Organ weights: epididymes, testes, ovaries

- Gross pathology: epididymes, testes, prostate gland incl. coagulating glands, ovaries, uterus, vagina, mammary gland.

Findings were considered to be within the range of normal background lesions. Only findings common in rats of this strain and age have been noted during gross pathology. All of them were within the range of historical control and are reflecting the usual individual variability. No significant findings for organ weights.

Overall, no indication of a reproductive toxic potential exist for the registration substance, neither from a reproductive screening study according to OECD 421 nor from additional data on reproductive organs from a repeated dose toxicity study. The NOAEL of 1000 mg/kg body weight with regard to developmental toxicity was established at the highest dose tested.

Justification for selection of Effect on fertility via oral route:
In order to assess the effects of the test substance on fertility via oral route, a valid OECD 421 study from 2011 in rats is available. Further a subacute 28-day oral gavage study in rats provides some information on effects on reproductive organs.

Effects on developmental toxicity

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

Kimisch 1 rated experimental study

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

A Klimisch-1-rated OECD 421 reproduction/development toxicity screening study has been conducted in the year 2011 on the registration substance. The substance proved to be not reprotoxic up to a dose level of 1000 mg/kg bw/day. Viabilitiy index was 100% in the control group and at the dose levels 300 and 1000 mg/kg bw/day and 94.5% at the dose level of 100 mg/kg bw/day. This difference in viability index in the 100 mg/kg bw/day was due to five pups which were missing on day 2 post partum. All these pups were from one litter. No further posnatal loss was noted at any dose level. No findings were noted in pups at first litter check or during the first 4 days post partum. Pups sex ratio was not affected by the treatment with the test item at any dose level. No effects were observed on pup body weight and body weight gain during the first four days post partum at any dose level. No findings were noted during macroscopic examination of pups at any dose level.

Justification for selection of Effect on developmental toxicity: via oral route:
In order to assess the effects of the test substance on development toxicity via oral route, a valid OECD 421 study from 2011 in rats is available. No other studies are available.

Justification for classification or non-classification

The registered substance proved to be not toxic on reproduction up to a dose level of 1000 mg/kg bw/d, thus no classification of the test item is required.