D-Glucitol, 1-deoxy-1-(methylamino)-, N-[C8-16 (even numbered) and C18 unsaturated acyl] derivs.

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

1991

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study well documented, followed guideline with minor deviation, GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1991

Report date:

1991

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

yes

Remarks:

(Animals were treated during organogenesis phase Day 6-15 of gestation instead of guideline recommendation of dosing through the day of implantation to one day prior to scheduled kill)

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Glucamide 24

IUPAC Name:

Glucamide 24

Constituent 2

Reference substance name:

287735-50-6

Cas Number:

287735-50-6

IUPAC Name:

287735-50-6

Test material form:

other: solid

Details on test material:

-Name of test material: C12/14 GS Base
-TSIN: SS0001.01
-Substance type: Other
-Physical state: White opaque solid gel
-Stability under test conditions: The test substance vehicle mixture contained the required amount of test substance 7 days following its preparation
-Storage condition of test material: Room temperature

Test animals

Species:

rat

Strain:

other: Crl:CD VAF/Plus®

Details on test animals or test system and environmental conditions:

TEST ANIMALS
-Source: Sprague-Dawley derived Crl:CD® VAF/Plus rats were obtained from Charles River Laboratories, Portage, Michigan
-Age at study initiation: 12.5 weeks (sexually mature virgin females)
-Weight at study initiation (on Day 0 of gestation): 221 – 227 g
-Housing: The females were housed individually in suspended, stainless steel wire-mesh cages from receipt until euthanasia except during mating.
-Diet: Certified Rodent Chow # 5002; ad libitum
-Water: Tap water, ad libitum
-Acclimation period: 10 days

ENVIRONMENTAL CONDITIONS
-Temperature: 72 – 73°F (mean temperature ± SD: 73 ± 0.4°F)
-Relative humidity: 39 – 50% (mean relative humidity ± SD: 42 ± 5.4%)
-Air changes: Not reported
-Photoperiod: 12 hours of fluorescent light per day

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Remarks:

deionized

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: The required amount of test substance for each group was dissolved into vehicle by heating on water bath at 50°C. Additional vehicle was added to obtain final volume of test substance preparation and mixed using a magnetic stir bar and stir plate. The test substance preparation was dispensed into amber glass containers and stored refrigerated. All the dosing solution concentrations were adjusted for active ingredient (45% aqueous solid gel).
-Rate of preparation of dose formulation: Weekly

VEHICLE
-Concentration in vehicle: 1.5, 15 and 36.3 mg/mL for dose of 15, 150 and 363 mg/kg bw respectively
-Amount of vehicle: 10 mL/kg bw/day

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Stability and periodic analysis of test substance preparations (during Day 6 to 13) were carried out using HPLC method. The details of method are provided in the study report. The test preparation for use during study week 1 and 2 contained 95 to 102% of the desired test substance concentrations.

Details on mating procedure:

-Impregnation procedure: Cohoused
-M/F ratio per cage: 1:1
-Length of cohabitation: Not reported
-Verification of same strain and source of both sexes: Yes
-Proof of pregnancy: Presence of copulatory plug was taken as Day 0 of gestation.

Duration of treatment / exposure:

Gestation days 6-15 (10 days)

Frequency of treatment:

Once daily

Duration of test:

Approximately 25 days

Doses / concentrations

Remarks:

Doses / Concentrations:
0 (control), 15, 150 and 363 mg/kg bw/day
Basis:
actual ingested

No. of animals per sex per dose:

25 mated females per dose

Control animals:

yes, concurrent vehicle

Details on study design:

-Dose selection rationale: The dosages were selected by the study sponsor on the basis of data from a previously conducted study (IRDC 191-1501).
-Rationale for animal assignment: Mated females were consecutively assigned in block design to one control and 3 treatment groups. The order in which mated females were assigned corresponded to the day the copulatory plug was observed and the order in which the animal appeared on the breeding record.
-Rationale for test system: The rat is acceptable model for developmental toxicity studies. The laboratory has historical control data on the incidence of parameters in this strain from this source. This strain is susceptible to known developmental toxicants.
-Dose volume: 10 mL/kg bw

Examinations

Maternal examinations:

MORTALITY AND SIGNS OF OVERT TOXICITY: Yes
-Time schedule: Twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
-Time schedule: At least once daily from Day 6 to 20 of gestation

BODY WEIGHT: Yes
-Time schedule: The individual maternal body weights were recorded on gestation Days 0, 6, 9, 12, 16 and 20. Body weights of non gravid animals were recorded but were not included in the mean calculations.

FOOD CONSUMPTION: No

WATER CONSUMPTION: No

POST-MORTEM EXAMINATIONS: Yes
-Sacrifice on gestation Day # 20; all animals were euthanized by CO2 inhalation.
-Organs examined: The abdominal and thoracic cavities and organs of females were examined for gross morphological changes.

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes, the uterus was excised and gravid uterine weights were recorded.
Examinations included:
-Number of corpora lutea: Yes
-Number of implantations: Yes
-Location of viable and non viable fetus: Yes
-Number of live fetuses: Yes
-Early resorptions: Yes
-Late resorptions: Yes
-Other: Uteri from females that appeared non gravid were opened and placed in 10% ammonium sulphide solution for detection of implantation.

Fetal examinations:

-External examinations: Yes, individual fetuses were weighed, sexed, tagged and examined for external malformations and variations.
-Soft tissue examinations: Yes, about 1/2 of the fetuses were placed in Bouin’s solution for subsequent soft tissue examination using Wilson’s razor blade sectioning techniques.
-Skeletal examinations: Yes, about 1/2 of the fetuses were fixed in ethanol, macerated with KOH, stained with Alizarin Red S and cleared with glycerin for subsequent skeletal examinations. Fetal findings were classified as malformations or developmental variations.
-Head examinations: No

Statistics:

All statistical analysis compared the treatment groups with the control with the level of significance at p≤0.05 and p≤0.01. All means were accompanied by standard deviations.
Mean maternal body weight, body weight changes, mean number of corpora lutea, total implantations, live fetuses, gravid uterine weight and mean fetal body weights were compared by one-way analysis of variance, Bartlett’s test for homogeneity of variance, and the appropriate t-test. Significance of difference was determined by Dunnet’s multiple comparison tables or pair wise comparison with a Bonferroni’s correction.
Male to female sex ratios and proportions of litters with malformations and developmental variations were compare using Chi-square test or Fisher’s exact probability test to determine the significance of difference.
The proportions of resorbed and dead fetuses and post-implantation losses were compared by the Mann-Whitney U-test to determine the significance of difference.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Maternal toxic effects:yes. Remark: (at 363 mg/kg body weight)

Details on maternal toxic effects:
MORTALITY: No mortality was observed during the study.

CLINICAL OBSERVATIONS: Presence of test material around the nose and/or mouth was observed at the high dose level in four animals. Test material around the mouth was observed in a single animal at mid dose level but was not considered to be treatment related. Post dose pharmacological signs of increased salivation in all animals and decreased activity in two animals were observed in the high dose group. Increased salivation was also observed in low incidence in themid dose group.

MATERNAL BODY WEIGHTS AND BODY WEIGHT CHANGES:
A statistically and biologically significant inhibition of body weight gain was observed in the high dose group relative to the control group during the first treatment interval of gestation Days 6–9 and during the overall treatment period of gestation Days 0–20.
A slight inhibition of maternal body weight gain was observed for the treatment interval of gestation Days 6–9 and Days 12–16 in the high dose group when compared with the control group.
A slight increase in maternal body weight gain was observed for treatment interval of gestation Days 16-20 at the high dose level when compared with the control group. This increase was considered to be compensatory for the earlier inhibition and therefore considered to be related to treatment.

NECROPSY FINDINGS: No treatment related differences were noted at necropsy; all findings were observed in low incidence.

CESAREAN SECTION EXAMINATION:
No treatment related differences were observed with regards to cesarean section parameters; values in the treated group were generally comparable with those of control group.

Effect levels (maternal animals)

1

Dose descriptor:

NOAEL

Effect level:

150 mg/kg bw/day

Based on:

act. ingr.

Basis for effect level:

other: maternal toxicity

2

Dose descriptor:

NOAEL

Effect level:

>= 363 mg/kg bw/day

Based on:

act. ingr.

Basis for effect level:

other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
MALFORMATIONS
No biologically meaningful difference was observed between the incidence of fetal malformations in the treated groups and that of control groups. The malformation occurred were primarily in single incidences were sporadically dispersed among the control and treated groups. No dose related trend was observed with regard to malformations.

DEVELOPMENTAL VARIATIONS:
The incidence and type of developmental variations observed among fetuses from treated group were generally comparable with those of the control group, or observed in sporadic instances with no apparent dose related trend. There was a slight increase in incidence of fetuses observed with unossified sternabrae in the mid and high-dose groups, however, no dose related trend was observed. The variation observed were considered to be due to normal biological variability and not related to treatment.

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

Administration of the test substance to female rats by oral gavage during Days 6-15 of gestation at dose levels of 0, 15, 150 and 363 mg/kg bw/day resulted in a NOAEL of ≥ 363 mg/kg bw for developmental toxicity.

Executive summary:

The developmental toxicity study of Glucamide 24 was conducted following methods comparable to the OECD Guideline 414 (Prenatal Developmental Toxicity Study).

Sexually mature female Crl:CD VAF/Plus® rats (Source: Charles River Laboratories, Portage, Michigan), weighing 221–227 g were used in this study. After receipt, animals were acclimated for 10 days and maintained under standard laboratory conditions (temperature: 7 –73°F, relative humidity: 39 – 50%, photoperiod: 12 hrs dark / 12 hrs fluorescent light). The animals were housed individually in suspended, stainless steel wire-mesh cages from receipt until euthanasia except during mating. The animals were fed on Certified Rodent Chow # 5002 and tap water, ad libitum.

One male and one female rat of same source and same strain were placed together for mating. The presence of a copulatory plug was taken as Day 0 of gestation. Mated females were consecutively assigned to the following treatment groups of 25 females each:

0 (vehicle control), 15, 150 and 363 mg/kg bw

Deionized water served as the vehicle control. The test substance was administered to mated female rats by oral gavage once daily beginning from Day 6 through to Day 15 of gestation. All animals were observed twice daily for mortality and at least once daily for detailed clinical observations. Body weights were recorded on Days 0, 6, 9, 12, 16, and 20 of gestation.

On Day 20 of gestation, animals were euthanized and cesarean section examinations were performed. The ovaries and uterine content (number of corpora lutea, number of implantations, and numbers of early and late resorptions, number and distribution of implantation sites) were examined after sacrifice.

Individual fetuses were weighed, sexed, tagged and examined for external malformations and variations. About half of all fetuses were examined for soft tissue changes using Wilson’s razor blade sectioning techniques. The other half of fetuses were fixed in ethanol, macerated with KOH, stained with Alizarin Red S and cleared with glycerin for subsequent skeletal examinations. Fetal findings were classified as malformations or developmental variations.

No mortality was observed during the study. No treatment related differences were noted at necropsy; all findings were observed in low incidence.

No treatment related differences were observed with regards to cesarean section parameters; values in the treated group were generally comparable with those of control group.

Maternal toxicity manifested as test material around the nose and/or mouth, post dose increased salivation and decreased activity, and significant inhibition of maternal body weight gain at the 363 mg/kg bw dose level.

The incidence of malformations and variations observed among fetuses in the treated group were generally comparable with those of the control group. The observations were sporadic with no apparent dose related trend. Therefore, no apparent developmental toxicity was observed at any dose level evaluated in the study.

Based on above, administration of Glucamide 24to female Crl:CD VAF/Plus rats by oral gavage during Days 6-15 of gestation at dose levels of 0, 15, 150 and 363 mg/kg bw/day resulted in a NOAEL of ≥ 363 mg/kg bw for developmental toxicity

The NOAEL for maternal toxicity was established at 150 mg/kg bw (based on clinical observations and body weight).

This developmental toxicity study is acceptable and satisfies the OECD Guideline 414 requirement.