Dimolybdenum trizinc nonaoxide

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

A reliable OECD 422 study is available conducted in accordance with GLP. In this study no effects were observed on reproductive parameters or in offspring and the NOAEL was set as the highest dose tested.

Link to relevant study records

Reference

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Reason / purpose for cross-reference:

reference to same study

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Wistar

Remarks:

RccHan™;WIST

Details on species / strain selection:

Standard for use in regulatory toxicology studies

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Envigo RMS Limited.
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: Males 84 to 90 days and females 98 to 104 days
- Weight at study initiation: Males 319 to 355g and females 195 to 237g
- Fasting period before study: No
- Housing: Cages comprised of a polycarbonate body with a stainless steel mesh lid; changed at appropriate intervals. Solid (polycarbonate) bottom cages were used (except during pairing). Grid bottomed polypropylene cages were used during pairing. These were suspended above absorbent paper which was changed daily during pairing.
- Diet (e.g. ad libitum): SDS VRF1 Certified pelleted diet provided ad libitum (removed overnight before blood sampling for hematology and blood chemistry investigations).
- Water (e.g. ad libitum): Potable water from the public supply via polycarbonate bottles with sipper tubes, non restricted.
- Acclimation period: Six days before commencement of treatment (males) and 20 days before commencement of treatment (females).

DETAILS OF FOOD AND WATER QUALITY: The diet contained no added antibiotic or other chemotherapeutic or prophylactic agent. Certificates of analysis for the diet are scrutinized and approved before any batch of diet was released for use. Certificates of analysis were routinely provided by the water supplier.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): Between 20 to 24°C
- Humidity (%): Between 40 and 70%
- Air changes (per hr):
- Photoperiod (hrs dark / hrs light): Artificial lighting, 12 hour light/dark cycle.

IN-LIFE DATES: From: To: 30 July 2019 (estrous cycle evaluation) to 13 October 2019 (last female necropsy).

Route of administration:

oral: gavage

Vehicle:

water

Remarks:

Purified water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
Starting with the lowest concentration, a sufficient amount of vehicle was added to the pre weighed test item to obtain a solution. Any large particles were crushed using a spatula and the solution was mixed with a magnetic stirrer. Once fully dissolved, the suspension was transferred to a measuring cylinder and made to the required volume using the vehicle. The suspension was transferred to a suitable container and stirred again using a magnetic stirrer. Formulations were transferred to their final containers using a syringe, whilst magnetically stirred. Since the formulation analysis method was not stability indicating, formulations were prepared daily and administered within four hours of preparation.

VEHICLE
- Justification for use and choice of vehicle (if other than water): Purified water
- Concentration in vehicle: 0, 6, 12 and 25 mg/mL
- Amount of vehicle (if gavage): 5 mL/kg/day

Details on mating procedure:

- M/F ratio per cage: 1:1
- Length of cohabitation: Up to two weeks
- Proof of pregnancy: Ejected copulation plugs in cage tray and sperm in the vaginal smear. Day 0 of gestation counted as day evidence of mating was detected.
- Further matings after two unsuccessful attempts: No
- After successful mating each pregnant female was caged (how): Individually
- Any other deviations from standard protocol: None

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

As per repeated dose entry.

Duration of treatment / exposure:

Males: Two weeks before pairing up to necropsy after a minimum of five weeks of treatment.
Females: Two weeks before pairing, then throughout pairing and gestation until Day 13 of lactation.

Frequency of treatment:

Daily via oral gavage

Details on study schedule:

- Age at mating of the mated animals in the study: Males 84 to 90 days and females 98 to 104 days

Dose / conc.:

0 mg/kg bw/day (nominal)

Dose / conc.:

30 mg/kg bw/day (nominal)

Dose / conc.:

60 mg/kg bw/day (nominal)

Dose / conc.:

125 mg/kg bw/day (nominal)

No. of animals per sex per dose:

10 males and females per dose

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: The dose levels selected for investigation in this OECD 422 study (0, 30, 60 and 125 mg/kg/day) were based on the results of a 14-day preliminary study. In the preliminary study dose levels of 62.5, 250, 500 and 1000 mg/kg/day were investigated in male and female Han Wistar rats. Treatment at doses of 250, 500 and 1000 mg/kg/day were not tolerated, with all groups terminated early as a result of excessive body weight loss. Treatment at 62.5 mg/kg/day which showed no obvious effect on body weight or food consumption. Clinical signs of animals treated at 62.5 mg/kg/day did not indicate any systemic effect of treatment, and necropsy showed no abnormalities. Dose levels of 30, 60 and 125 mg/kg/day (follow the OECD guidelines with a regular interval) were considered suitable for this main investigation.

- Rationale for animal assignment (if not random): Estrous cycles were evaluated pre-treatment. After 14 days evaluation, animals that failed to exhibit typical 4-5 days cycles were not allocated to the study. On Day 1 of study all animals were weighed and body weights were reviewed before dosing commenced by Study Management. Body weight of animals did not exceed +/- 20% of the mean for each sex.

- Fasting period before blood sampling for clinical biochemistry: Yes. Food witheld the night before.

Positive control:

Not a guideline requirement

Parental animals: Observations and examinations:

See repeated dose study for details on parental animal observations and examinations.

Oestrous cyclicity (parental animals):

Smears were taken for 15 days before pairing using cotton swabs (dry smears). Wet smears were taken for 14 days before treatment, after pairing until mating and for 4 days before scheduled termination (nominally Days 10-13 of lactation).

Sperm parameters (parental animals):

Not examined

Litter observations:

STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: No

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
Clinical observations (examined approximately 24 hours after birth and then daily), litter size (daily), sex ratio (Days 1, 4, 7, 11 and 13), individual bodyweights (Days 1, 4, 7, 11 and 13), ano-genital distance (day 1) and nipple/areolae count (Day 13 in males).

GROSS EXAMINATION OF DEAD PUPS:
All animals were subject to an external macroscopic examination; particular attention was paid to the external genitalia. Animals observed with external abnormalities were retained pending possible future examination. Thyroid glands were preserved from one male and one female in each litter.

Postmortem examinations (parental animals):

See repeated dose study for details on parental animal observations and examinations.

Postmortem examinations (offspring):

GROSS EXAMINATION OF DEAD PUPS:
All animals were subject to an external macroscopic examination; particular attention was paid to the external genitalia. Animals observed with external abnormalities were retained pending possible future examination. Thyroid glands were preserved from one male and one female in each litter.

Reproductive indices:

Pre-coital interval, percentage mating, conception rate, fertility index, gestation length and index, litter size and post-implantation survival index,

Offspring viability indices:

Survival index, live birth index, viability index, lactation index and sex ratio.

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

See repeated dose entry for paternal data.

Dermal irritation (if dermal study):

not examined

Mortality:

mortality observed, non-treatment-related

Description (incidence):

See repeated dose entry for paternal data.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

See repeated dose entry for paternal data.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

See repeated dose entry for paternal data.

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

See repeated dose entry for paternal data.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

See repeated dose entry for paternal data.

Urinalysis findings:

not examined

Behaviour (functional findings):

effects observed, non-treatment-related

Description (incidence and severity):

See repeated dose entry for paternal data.

Immunological findings:

no effects observed

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

See repeated dose entry for paternal data.

Histopathological findings: neoplastic:

not examined

Other effects:

not examined

Reproductive function: oestrous cycle:

no effects observed

Description (incidence and severity):

Estrous cycles were considered unaffected by treatment when compared with controls. All females showed regular 4 or 5 day cycles prior to treatment and there was no effect of treatment of estrous cycle regularity. All females mated within 1-4 days with the exception of one Control female which was observed to be acyclic during pairing. At termination, all females were observed to be in diestrus.

Reproductive function: sperm measures:

not examined

Reproductive performance:

no effects observed

Description (incidence and severity):

Pre-coital interval, mating performance, gestation length, gestation index and fertility were considered unaffected by treatment with the substance, when compared to Controls.

Dose descriptor:

NOAEL

Effect level:

125 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Remarks on result:

other: No effects observed on reproduction parameters. NOAEL is set as the high dose.

Clinical signs:

no effects observed

Description (incidence and severity):

There were no treatment related clinical signs observed amongst offspring following parental administration of the test substance.

Dermal irritation (if dermal study):

not examined

Mortality / viability:

no mortality observed

Description (incidence and severity):

There was no effect of parental treatment of the mean number of offspring survival indices.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Body weights of male and female offspring on Day 1 of age and subsequent body weight gain until Day 13 of age was similar to that of the Control in all treated groups. Minor intergroup differences were considered to reflect the marginally higher litter sizes in the 30 or 125 mg/kg/day groups.

Food consumption and compound intake (if feeding study):

not examined

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

Sexual maturation:

not examined

Anogenital distance (AGD):

no effects observed

Description (incidence and severity):

There was no effect of treatment on ano-genital distance in the male and female offspring at any dose level investigated.

Nipple retention in male pups:

no effects observed

Description (incidence and severity):

No nipples were observed in male offspring of parents treated with the test substance.

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Description (incidence and severity):

Macroscopic examination of offspring that either died prematurely or at scheduled termination on Day 13 of age did not reveal any findings that could be attributed to treatment.

Histopathological findings:

not examined

Other effects:

not examined

Behaviour (functional findings):

not examined

Developmental immunotoxicity:

not examined

The estrous cycle investigations, during the pre-mating period was unaffected, and all animals with litters were in di-estrus at termination, indicating that estrous cycles had ceased during lactation. Pre-coital interval, mating performance, gestation length and gestation index were unaffected by treatment. The litter size, sex ratio, survival, ano-genital distance and growth of the offspring (F1) were unaffected by treatment, the males did not develop nipples and there were no parental treatment related macroscopic findings in the offspring. There was no effect on serum T4 concentrations in adult males and females treated at 30, 60 or 125 mg/kg/day, nor in the F1 offspring.

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

125 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Remarks on result:

other: No effects observed relating to offspring. NOAEL is set as the high dose.

Reproductive effects observed:

no

Conclusions:

General systemic toxic potential in Han Wistar rats, including a screen for reproductive/developmental effects and assessment of endocrine disruptor relevant endpoints, were evaluated with by oral gavage administration; concentrations of 30, 60 or 125 mg/kg/day were generally well-tolerated when administered daily for at least five weeks in males, and in females for two weeks before pairing, throughout pairing and gestation to Day 13 of lactation.

As administration of the substance at doses up to and including 125 mg/kg/day did not affect the reproductive performance of male and females, or the development of their offspring, it was concluded that the NOAEL for reproductive/developmental toxicity was the high dose of 125 mg/kg/day.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

125 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

Reliable OECD 422 study available conducted to GLP.

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Effects on developmental toxicity

Description of key information

A reliable OECD 422 study is available conducted in accordance with GLP. In this study no effects were observed on reproductive parameters or in offspring and the NOAEL was set as the highest dose tested.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

125 mg/kg bw/day

Study duration:

subacute

Species:

rabbit

Quality of whole database:

Reliable OECD 422 study available conducted in accordance with GLP

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Justification for classification or non-classification

The oral administration of the test substance to rats of both sexes by gavage did not result in any adverse treatment-related effects to reproductive or developmental endpoints. The overall NOAEL for the study was set as the highest dose tested (125 mg/kg/day). As no adverse effects were observed, classification for reproductive toxicity in accordance with EU CLP (1272/2008, as amended) or UN GHS is not warranted.