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Toxicological information

Toxicity to reproduction

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Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
23.10.1991 to 26.05.1992
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study
Cross-reference
Reason / purpose for cross-reference:
reference to same study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1992
Report date:
1992

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Combined Repeated Dose and Reproductive / Developmental Toxicity Screening Test (Precursor Protocol of GL 422)
Version / remarks:
draft 22 March 1990
GLP compliance:
no
Remarks:
The report states that the study was conducted according to the principles of GLP, but the work was not certified and was not subject to Quality Assurance review .
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
2,2,2',2'-tetrakis(hydroxymethyl)-3,3'-oxydipropan-1-ol
EC Number:
204-794-1
EC Name:
2,2,2',2'-tetrakis(hydroxymethyl)-3,3'-oxydipropan-1-ol
Cas Number:
126-58-9
Molecular formula:
C10H22O7
IUPAC Name:
2,2'-[oxybis(methylene)]bis[2-(hydroxymethyl)propane-1,3-diol]
Specific details on test material used for the study:
- Name of test material (as cited in study report): dipentaerythritol
- Substance type: white powder
- Physical state: solid
- Analytical purity: 96.2%
- Purity test date: received 2 September 1991
- Lot/batch No.: 9404
- Expiration date of the lot/batch: 2 November 1992
- Storage condition of test material: within the Formulation Department at room temperature in darkness

Test animals

Species:
rat
Strain:
other: Crl:CD (SD) BR VAF/Plus
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River UK Ltd.
- Age at study initiation: (P) Males 12 wks ±1 day, Females (sexually mature, virgin) 7 weeks ±1day;
- Weight at study initiation: (P) Males: 440-446 g; Females: 224-228 g
- Fasting period before study: overnight
- Housing: pre-mating; suspended stainless steel cages equipped with solid sides and wire grid front, back, floor, and top. During mating male and female pairs were housed in plastic breeding cages (North Kent Plastics, RM2 type).
- Acclimatisation period: 1 week
- Diet : Biosure Laboratory Animal Diet No. 1 ad libitum
- Water : tap water ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21
- Humidity (%): 55
- Air changes (per hr): not specified
- Photoperiod (hrs dark / hrs light): 12/12

Five males and five females were supplied for health check purposed. They were killed within 24 hours of arrival and subjected to routine macroscopic examination.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: 1% methylcellulose
Details on exposure:
The test material was ground in a mortar with a small volume of 1% methylcellulose until a smooth paste was formed. The formulation was then gradually made up to volume and mixed using a high speed homogeniser. A series of suspensions were then made to give the required concentrations. Formulations were prepared daily and dosed on the day of preparation.
Details on mating procedure:
Rats were paired 1 male to 1 female during the mating period (14 days). Mating was confirmed (by the presence of a sperm plug and vaginal smears). After the mating period, the males and females were returned to standard single housing.
Analytical verification of doses or concentrations:
no
Details on analytical verification of doses or concentrations:
Analytical verification of doses was not carried out.
Duration of treatment / exposure:
Males: 7 weeks; Females: 9 weeks.
Frequency of treatment:
Daily
Details on study schedule:
Rats underwent 2 weeks of treatment prior to blood collection and commencement of the mating phase. The mating phase lasted for 2 weeks. Repeat blood collections were taken from female rats prior to parturition, at week 5. Males were sacrificed after 7 weeks of treatment. Females were sacrificed after 9 weeks of treatment of post partum day 4 (non-pregnant females were sacrificed after 9 weeks treatment).
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Dose / conc.:
500 mg/kg bw/day (actual dose received)
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
20 per dose, 10 males, 10 females
Control animals:
yes
Details on study design:
A range finding study was conducted prior to the main study, the limit dose of 1000 mg/kg/day was established as a suitable high dosage level. Controls were concurrent but it is not stated whether they were unexposed, or administered the vehicle alone.
Positive control:
A positive control was not examined.

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily


BODY WEIGHT: Yes
- Time schedule for examinations: recorded at allocation to groups, commencement of treatment and thereafter weekly. Females also weighed daily from the commencement of the mating period until parturition and Days 0 and 4 post partum.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes, measured during the pre-treatment and pre-mate period for males and females, during the post mate period for males and post partum for females. Water intake was only measured by visual observation, and could therefore not be subjected to statistical analysis.


HAEMATOLOGY: Yes, assessed after two weeks of treatment, and again after 5 weeks treatment in females
- Anaesthetic used for blood collection: Light ether anaethesia
- Animals fasted: Yes, overnight
- Parameters examined: PCV, haemoglobin, RBC, MCHC, MCV, total and differential WBC, platelets, examination for abnormal cell morphology, thrombotest, reticulocyte count.


CLINICAL CHEMISTRY: Yes, assessed after two weeks of treatment, and again after 5 weeks treatment in females.
- Animals fasted: Yes, overnight
- Parameters examined: Glucose, GPT, GOT, CPK, gammaGT, total protein albumin, globulin, urea nitrogen, AP, total bilirubin, creatine, sodium, potassium, calcium, inorganic phosphorus, chloride, cholesterol.

Oestrous cyclicity (parental animals):
Vaginal smears were taken daily starting from 1 week prior to mating and throughout the mating phase.
Sperm parameters (parental animals):
Sperm parameters were not examined.
Litter observations:
For each litter the pups were counted, sexed, weighed and examined for gross abnormalities as soon as possible after birth. Litters were examined daily for dead/missing (i.e. cannibalised) pups. Pups were weighed again on Day 4 post partum.

Litter weight and mean pup weight were calculated from individual pup weight. Sex ratios were calculated at birth and at day 4.
Pup loss on completion of parturition was calculated according the formula: ((total no. young at birth - no. live young) / total no. young at birth) x100
Cumulative pup loss was calculated according to the formula: ((total no. young at birth - no. live young at Day 4) / total no. young at birth) x100.
Postmortem examinations (parental animals):
All rats were subject to gross necropsy at study termination, and examined for macroscopic abnormalities. Organ weights: adrenals, brain, epididymides, heart, kidneys, liver, lungs, ovaries, prostrate, testes, thymus, spleen. The following tissues were examined histologically (* denotes tissues examined from control and high dose groups only): adrenals*, brain*, epididymides*, heart*, kidneys*, liver*, lungs, mammary gland, macroscopically abnormal tissues, ovaries*, pituitary*, prostate*, seminal vesicles*, spleen*, testes*, thymus, thyroids, uterus* and vagina*.
Implantation loss was calculated according to the formula: ((no. implantation sites - total young born) / no. implantation sites) x 100.
Postmortem examinations (offspring):
Litters were examined for abnormalities and sex confirmed.
Statistics:
ANOVA followed by Williams' test, or Kruskal-Wallis 'H' statistic followed by Shirley's test where appropriate.
Reproductive indices:
Fertility index = (no. pregnant females / no. females showing evidence of copulation) x100.
Gestation index = (no. pregnant females with live young / no. pregnant females) x 100.
Offspring viability indices:
Viability index (day 4) = (no. live young at day 4 / no. live young at birth) x100.

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
no effects observed
Description (incidence and severity):
For more details, see the " details on results (P0) " section below.
Mortality:
no mortality observed
Description (incidence):
For more details, see the " details on results (P0) " section below.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
For more details, see the " details on results (P0) " section below.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
For more details, see the " details on results (P0) " section below.
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
For more details, see the " details on results (P0) " section below.

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
no effects observed
Description (incidence and severity):
For more details, see the " details on results (P0) " section below.
Reproductive function: sperm measures:
not examined
Reproductive performance:
no effects observed
Description (incidence and severity):
For more details, see the " details on results (P0) " section below.

Details on results (P0)

There were no deaths during the study. There were no clinical signs associated with treatment. The bodyweight gain of treated males was similar to that of controls throughout the study. Females given 1000 mg/kg/day showed reduced gain during pregnancy but recovered post partum.
No effects of treatment on food intake were apparent over the time periods of measurement.
There were no changes in the haematological and biochemical parameters measured that were clearly indicative of a reaction to treatment.
There were no differences in organ weights that indicated any treatment-related effects.
There were no differences or changes noted at histopathological examination that indicated any treatment-related effects.
The oestrus cycle was unaffected by treatment and the pregnancy rate was good in all groups. Time to mating and duration of pregnancy were within normal limits. Two females were not pregnant, and one dam exhibited total resorption of her litter.

Effect levels (P0)

Key result
Dose descriptor:
NOAEL
Effect level:
> 1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No effects were seen

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Description (incidence and severity):
For more details, see the " details on results (F1) " section below.
Mortality / viability:
mortality observed, non-treatment-related
Description (incidence and severity):
For more details, see the " details on results (F1) " section below.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
For more details, see the " details on results (F1) " section below.
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Description (incidence and severity):
For more details, see the " details on results (F1) " section below.
Histopathological findings:
not examined

Details on results (F1)

There were no obvious effects of treatment on litter size or sex ratio at birth, or on survival at post partum day 4. Litter and mean pup weight also appeared unaffected. There were no findings at necropsy, and no clinical signs were reported. A small number of pups died at birth, or by post partum day 4, none of these deaths were attributed to treatment. Litter data are shown in Table 1.

Effect levels (F1)

Dose descriptor:
NOAEL
Generation:
F1
Effect level:
> 1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: There were no obvious effects of treatment on litter size or sex ratio at birth, or on survival at post partum day 4. Litter and mean pup weight also appeared unaffected. There were no findings at necropsy, and no clinical signs were reported.

Target system / organ toxicity (F1)

Critical effects observed:
no

Overall reproductive toxicity

Reproductive effects observed:
no

Any other information on results incl. tables

Table 1: Litter data

Group No. of Animals Implant sites Implant loss % At birth At day 4
Litter size Pup loss % Litter Wt (g) Mean pup wt. (g) Litter size Cumulative loss% Litter wt (g) Mean pup wt.(g)
Mated Pregnant male female Total %males live male female Total %males
Control 10 10 17.5 4.6 9 7.7 16.7 54.9 16.3 2.3 102.9 6.3 8.7 7.3 16 55.3 4.2 161.6 10.2
500 mg/kg/day 10 8 18.4 4.1 9.4 8.3 17.6 53.4 17.5 0.7 107.8 6.2 9.3 8 17.3 53.8 2.1 165.2 9.6
1000 mg/kg/day 10 9 17.1 3.7 7.7 8.8 16.4 46.2 16.4 0 101.4 6.2 7.3 8.6 15.9 45.8 3.4 158.2 10

Applicant's summary and conclusion

Conclusions:
The study author concluded that there were no findings at either of the dosage levels employed in this study to indicate an immediate requirement for more detailed studies.
Executive summary:

The study was conducted to assess the repeated dose oral toxicity and toxicity to reproduction of dipentaerythritol, in accordance with OECD guideline 422 (draft version; 22 March 1990).

Sexually mature rats were exposed to dipentaerythritol via oral gavage at dose levels 0, 500, and 1000 mg/kg bw/d. The rats were exposed throughout pregnancy and the effects on both parents and the F1 generation were assessed.

Male rats were exposed for 7 weeks prior to sacrifice and necropsy. Females were exposed for 9 weeks until post partum day 4, when they along with their pups were sacrificed and subjected to gross necropsy.

There were no findings at either of the dosage levels in this study to indicate an immediate requirement for more detailed studies. No effects of treatment were observed on food intake, bodyweight change, haematology, biochemistry, organ weights, post mortem findings and histopathology. There did not appear to be any effects on mating or on pup viability.

The NOAEL for reproductive and developmental effects can therefore be considered to be 1000 mg/kg bw/d.