L-gluco-Octitol, 1,5-anhydro-6,8-dideoxy-, (7.xi.)-

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

02 - 26 November 2004

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP study conducted according to OECD Guideline 414 with minor deviation: cartilage structures were not submitted to evaluation due to a technical problem during the coloration (poor quality of water)

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:

2006

Report date:

2006

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

yes

Remarks:

cartilage structures were not submitted to evaluation due to a technical problem during the coloration (poor quality of water)

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Details on test material:

- Name of test material: see confidential details on test material
- Physical state: yellowish orange-colored liquid (on certificate), brown orange-colored liquid (at receipt)
- Lot/batch No.: see confidential details on test material
- Date of receipt: 5 August 2004
- Expiration date of the lot/batch: November 2006
- Storage condition of test material: Stored at at room temperature

Test animals

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories France, L’Arbresle, France
- Age at study initiation: 10 weeks
- Weight at study initiation: 224-308 g (mean: 263 g)
- Housing: Animals were housed individually in suspended wire-mesh cages.
- Diet: A04 C pelleted maintenance diet (SAFE, Epinay-sur-Orge, France) distributed weekly, ad libitum
- Water: Tap water (filtered with a 0.22 μm filter), ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature: 22 ± 2 °C
- Humidity: 50 ± 20 %
- Air changes: Approximately 12 cycles/h of filtered, non-recycled air
- Photoperiod: 12 h dark / 12 h light

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: 60/40 (w/w) propylene glycol/purified water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
- Test item was diluted with the required quantity of vehicle in order to achieve the desired concentrations and then homogenized using a magnetic stirrer.
- Test item dosage forms were prepared for up to 9 days of use and stored at +4 °C. They were used each day within a maximum of 6 h storage at room temperature.

VEHICLE
- Concentration in vehicle: 20, 60 and 200 mg/mL
- Amount of vehicle (if gavage): 5 mL/kg bw/day
- Lot/batch no. of propylene glycol: S21645-174
- Source of propylene glycol: Aldrich (Saint-Quentin-Fallavier, France)

STABILITY:
- Two dosage forms were prepared at 10 and 200 mg/mL of test item to cover all the concentrations intended for use in this study.
- To evaluate the stability, each dosage form was sampled (in duplicate) after 0 (just after preparation), 6 h storage at room temperature, then after 4 and 9 days storage at +4 °C. The aliquots sampled on Day 4 were analyzed immediately after sampling.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

- Concentration of samples taken from each dosage form (including the control) prepared for use on the first and last days of the dosing period was determined.
- Results of analysis: Satisfactory agreement was observed between the nominal and actual concentrations of the test item in the administered dosage forms prepared for the first and last days of dosing, since the deviations from nominal concentration were in an acceptable range of ± 9 %.

Details on mating procedure:

- Impregnation procedure: Cohoused; females in proestrus were placed overnight in the home cage of singly housed stock males.
- M/F ratio per cage: 1:1
- Length of cohabitation: Overnight
- Verification of same strain and source of both sexes: Yes
- Proof of pregnancy: Sperm in vaginal smear or a sperm plug in situ referred to as Day 0 of pregnancy
- Monitoring of estrous cycle: During the week of mating, the estrous cycle stage was determined periodically from a fresh vaginal lavage (stained with methylene blue).

Duration of treatment / exposure:

14 days (Days 6-19 post-coitum)

Frequency of treatment:

Once daily

Duration of test:

21 days (Days 0-20 post-coitum)

Doses / concentrations

Remarks:

Doses / Concentrations:
100, 300 and 1000 mg/kg bw/day
Basis:
actual ingested

No. of animals per sex per dose:

24 mated females/dose

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Dose levels were selected on the basis of the results of preliminary study of prenatal developmental toxicity study (Study No. 28463 RSR) in which the test item was given orally to Sprague-Dawley female rats from Days 6-19 post-coitum at dose-levels of 100, 300 and 1000 mg/kg bw/day. In absence of signs of toxicity at the high dose-level, 1000 mg/kg bw/day was selected as the highest dose-level.
- Rationale for animal assignment: Before Day 3 post-coitum, the animals were allocated to the groups according to a stratification procedure based on body weight recorded on Day 0 post-coitum, to ensure comparatively similar mean body weight among groups. A larger number of animals than necessary were paired to permit the selection and/or replacement of individuals before start of treatment.

Examinations

Maternal examinations:

CAGE SIDE & CLINICAL OBSERVATIONS: Yes
Time schedule:
- Mortality or signs of morbidity: At least twice a day
- Clinical signs: Once daily

BODY WEIGHT: Yes
- Time schedule for examinations: Days 0, 3, 6, 9, 12, 15, 18 and 20 post-coitum

FOOD CONSUMPTION: Yes
- Time schedule: Days 0-3, 3-6, 6-9, 9-12, 12-15, 15-18 and 18-20 post-coitum.

POST-MORTEM EXAMINATIONS: Yes
- Time schedule: On Day 20 post-coitum, females were sacrificed by asphyxiation using carbon dioxide followed by cervical dislocation and were subjected to a macroscopic post-mortem 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 and distribution of implantations: Yes
- Number and distribution of early and late resorptions: Yes
- Number and distribution of uterine scars: Yes
- Number and distribution of dead and live fetuses: Yes
- Gross evaluation of placentas: Yes

Fetal examinations:

- External examinations: Yes [all living foetuses]; weighed and examined for gender and external malformations including the observation of all visible structures, surfaces and orifices.
- Soft tissue examinations: Yes [half per litter]: fixed with Harrisson’s fluid and evaluated for detailed examination including the observation of all the organs and structures of the head, neck, thorax and abdomen.
- Skeletal examinations: Yes [all the remaining litters]; fixed with ethyl alcohol, stained with Alizarin Red S and examined for skeletal abnormalities including the observation of all the bone structures of the head, spine, rib cage, pelvis and limbs.

Statistics:

- All data are recorded using computerized software (Reprotox version B.1).
- Mean values were compared by one-way analysis of variance and Dunnett test (mean values being considered as normally distributed and variances being considered as homogeneous).
- Percentage values were compared by the Fisher exact probability test.

Indices:

- Pre-implantation loss: [(Number of corpora lutea - Number of implantation sites) / Number of corpora lutea] X 100
- Post-implantation loss: (Number of implantation sites - Number of live fetuses) / Number of implantation sites] X 100
- Fetal or litter incidence: Total number of fetuses or litters with a particular finding / Total number of fetuses or litters examined X 100
- Mean proportion of affected fetuses: Sum of proportion of fetuses affected in each litter / Total number of litters examined] X 100

Historical control data:

Historical control data was used to compare the incidences with control group.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Maternal toxic effects:no effects

Details on maternal toxic effects:
- No mortality and no clinical signs were observed.
- Body weight, food consumption and mean uterus weights were unaffected by treatment with the test item. The net body weight change was slightly lower at 1000 mg/kg bw/day but the difference from the control group was not statistically significant.
- No treatment-related necropsy findings were recorded.
- Pregnancy status of females: All females, except one in each of the 100 and 1000 mg/kg bw/day groups, were pregnant with live fetuses at scheduled sacrifice.

Effect levels (maternal animals)

Dose descriptor:

NOAEL

Effect level:

> 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
LITTER DATA:
- No dead fetuses were noted.
- Mean fetal body weights and mean number of resorptions recorded in the test item-treated groups were similar to those of the control group.

FETAL EVALUATION:
- No significant dose-dependent increase in external, visceral and skeletal malformations was induced by treatment with the test substance.
- At 1000 mg/kg bw/day, one fetus showed a cardiac malformation (ventricular septum defect) and one fetus from another litter presented a malformated left renal apparatus (marked dilated renal pelvis of the kidney and ureter). Since these were noted at a very low incidence, a relationship to treatment with the test item could be ruled out.
- When compared to the mean control values, both fetal incidence and litter incidences of unossification of the hyoid bone recorded in the 1000 mg/kg bw/day group were higher, and the mean values were above historical control data (see table 7.8.2/1). The incidence of this variation was close to the upper limit of historical data and the total number of affected fetuses in this group was statistically significant (17 vs. 5 in controls). However, in the absence of dose-related increase in incidence of this variation, a relationship to treatment with the test item was ruled out.

Effect levels (fetuses)

Dose descriptor:

NOAEL

Effect level:

> 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: fetotoxicity

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Any other information on results incl. tables

Table 7.8.2/1: Incidence of unossified hyoid bone

Dose-level (mg/kg bw/day)	0	100	300	1000
- fetal incidence (%), [0-9.5]	3	6.5	1.7	10.6
- litter incidence (%), [0-33.3]	20.8	21.7	8.3	47.8

[CIT historical data]

Applicant's summary and conclusion

Conclusions:

Under the test conditions, the No Observed Adverse Effect Level (NOAEL) of test item for maternal and fetal toxicity was considered to be greater than 1000 mg/kg bw/day in Sprague-Dawley rats.

Executive summary:

In a prenatal developmental toxicity study performed according to OECD Guideline 414 and in compliance with GLP, the test item was administered orally by gavage to groups of pregnant Sprague-Dawley, Crl CD® (SD) IGS BR rats (24/dose) at dose levels of 0 (vehicle), 100, 300 and 1000 mg/kg bw/day in 60/40 (w/w) propylene glycol/purified water from Days 6 to 19 post coitum. Clinical signs and mortality were checked daily. Body weight and food consumption were recorded at designated intervals. On Day 20 post-coitum, the dams were sacrificed and subjected to macroscopic examination. The gravid uterine weight, numbers of implantations, uterine scars, live and dead fetuses, early and late resorptions and corpora lutea were recorded. Fetuses were weighed and examined for external, soft tissue and skeletal malformations.

No mortality and no clinical signs were observed. Body weight and food consumption were unaffected by the treatment. No treatment-related necropsy findings were noted. The litter parameters in all groups were considered to be unaffected by the treatment. Fetal body weight measurement and external, soft tissue or skeletal examinations did not reveal any treatment-related effects.

Under the test conditions, the No Observed Adverse Effect Level (NOAEL) of test item for maternal and fetal toxicity was considered to be greater than 1000 mg/kg bw/day in Sprague-Dawley rats.