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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:
2008-09-25 to 2008-11-14
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Guideline study performed on a supporting substance (structural analogue). In accordance with the ECHA guidance document “Practical guide 6: How to report read-across and categories (March 2010)”, the reliability was changed from RL1 to RL2 to reflect the fact that this study was conducted on a read-across substance.
Cross-reference
Reason / purpose for cross-reference:
reference to same study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2009
Report date:
2009

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
other: OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Reference substance name:
Isostearic acid, esters with methyl α-D-glucoside
IUPAC Name:
Isostearic acid, esters with methyl α-D-glucoside
Constituent 2
Reference substance name:
138985-20-3
Cas Number:
138985-20-3
IUPAC Name:
138985-20-3
Test material form:
semi-solid (amorphous): gel
Remarks:
migrated information: paste

Test animals

Species:
rat
Strain:
other: Wistar Han (Crl:WI(Han))
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Age at study initiation (F0-treatment): approximately 10 weeks
- Weight at study initiation: male:279 - 317 g, female: 180 - 215 g
- Fasting period before study: no
- Housing:
Pre-mating: animals were housed in groups of 5 animals/sex/cage in Macrolon cages (MIII type, height 18 cm)
Mating: females were caged together with males on a one-to-one-basis in Macrolon cages (MIII type, height 18 cm).
Post-mating: Males were housed in their home cage (Macrolon cages, MIV type, height 18 cm) with a maximum of 5 animals/sex/cage. Females were individually housed in Macrolon cages (MIII type, height 18 cm).
Lactation: Offspring was kept with the dam until termination in Macrolon cages (MIII type, height 18 cm)
- Diet: ad libitum, pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany)
- Water: ad libitum, tap-water
- Acclimation period F0: at least 5 days prior to start of treatment


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 3°C (actual range: 18.4 to 22.2°C)
- Humidity (%): 30 - 70% (actual range: 37 - 94%)
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: 1% aqueous carboxymethyl cellulose
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
- Dose volume: 5 mL/kg bw. Actual dose volumes were calculated according to the latest body weight.

VEHICLE
- 1% aqueous carboxymethyl cellulose

TEST SUBSTANCE FORMULATION
- Stability of test substance in vehicle: stable in 1% aqueous carboxymethyl cellulose for at least 6 hours at room temperature over the concentration range 10 to 200 mg/mL (determined during this project).
- Method of formulation: formulations (w/w) were prepared daily, were homogenised to a visually acceptable level and dosed as soon as possible after preparation with a maximum of 2.5 hours after preparation. No adjustment was made for specific gravity of the test substance, vehicle, and/or
formulation. In order to obtain homogeneity, the test substance formulations were heated in a water bath with a maximum temperature of 45 °C for a maximum of 22 minutes. The test substance formulations were allowed to cool down to a temperature at a maximum of 40 °C prior to dosing.
Based on results of the thermal analysis performed by NOTOX (NOTOX project 488541), reaction and/or decomposition of the test substance were observed above approximately 175°C and, therefore, the test substance was considered to be stable at 45°C.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
- Analyses were performed on a single occasion after the treatment phase according to a validated method (NOTOX Project 488541).
- The accuracy of preparation was considered acceptable if the mean measured concentrations were 85-115%. Homogeneity was demonstrated if the coefficient of variation was <= 10%. Formulations were considered stable if the relative difference before and after storage was maximally 10%.
Details on mating procedure:
- M/F ratio per cage: 1/1
- Length of cohabitation: a minimum of 14 days of exposure
- Proof of pregnancy: sperm in vaginal lavage or by appearance of an intravaginal copulatory plug referred to as day 0 post coitum
- After successful mating each pregnant female was caged (how): Females were individually housed in Macrolon cages (MIII type, height 18 cm).
Duration of treatment / exposure:
Males: exposed for 30 days, i.e, 2 weeks prior to mating, during mating, and up to termination
Females: exposed for 42-44 days, i.e, during 2 weeks prior to mating, during mating, post-coitum, and during at least 4 days of lactation
Offspring: not treated
Frequency of treatment:
- Once daily for 7 days per week, approximately the same time each day with a maximum of 6 hours difference between the earliest and latest dose. Animals were dosed up to the day prior to scheduled necropsy.
Duration of test:
30/44 days
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 50, 150, 1000 mg/kg bw/day
Basis:
actual ingested
No. of animals per sex per dose:
F0 males: 10, F0 females: 10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Dose levels were based on results of the dose range finding study (NOTOX Project 488547, cited in this study under 138985-20-3_8.6.1_8.7.1_Evonik Goldschmidt_2009_OECD 422, Appendix 5)
- Rationale for animal assignment (if not random): This species and strain of rat has been recognized as appropriate for general and reproductive toxicity studies.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: at least twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: at least once daily
- Once prior to start of treatment and at weekly intervals this was also performed outside the home cage in a standard arena. Arena observations were not performed when the animals were mating, or housed individually.
- All symptoms were recorded and graded according to fixed scales:
Maximum grade 1: grade 0 = absent, grade 1 = present
Maximum grade 3 or 4: grade 1 = slight, grade 2 = moderate, grade 3 = severe, grade 4 = very severe


BODY WEIGHT: Yes
- Time schedule for examinations: first day of exposure and weekly thereafter; mated females were weighed on days 0, 4, 7, 11, 14, 17 and 20 post-coitum and during lactation on days 1 and 4


FOOD CONSUMPTION:
- Time schedule: weekly for males and females. It was not recorded during the breeding period. Food consumption of mated females was measured on Days 0, 4, 7, 11, 14, 17 and 20 postcoitum and during lactation on Days 1 and 4 post-partum


WATER CONSUMPTION:
- Subjective appraisal was maintained during the study, but no quantitative investigation introduced as no effect was suspected

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: immediately prior to scheduled post mortem examination, between 7.00 and 10.30 a.m.
- Anaesthetic used for blood collection: Yes (iso-flurane (Abbott Laboratories Ud., Zwolle, The Netherlands) anaesthesia)
- Animals fasted: Yes
- How many animals: from the first five mated males and the first five females with live offspring from each group
- Parameters: WBC, differential leucocyte count, RBC, Reticulocytes, Red blood cell distribution width (RDW), Haemaglobin, Haematocrit, MCV, MCH, MCHC, platelets, PT, APTT

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: immediately prior to scheduled post mortem examination, between 7.00 and 10.30 a.m.
- Animals fasted: Yes
- How many animals: from the first five mated males and the first five females with live offspring from each group
- Parameters: Alanine aminotransferase, Aspartate aminotransferase, Alkaline phosphatase,Total Protein, Albumin, Total Bilirubin, Urea, Creatinine,
Glucose, Cholesterol, Sodium, Potassium, Chloride, Calcium, Inorganic Phosphate

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: The assigned males were tested during Week 4 of treatment and the assigned females were tested during lactation (all before blood sampling)
- Dose groups that were examined: In the first five mated males and the first five females with live offspring, from each group
- Battery of functions tested: hearing ability, pupillary reflex, static righting reflex and grip strength (Score 0 =normal/present, score 1 =abnormal/absent); motor activity test (recording period: 12 hours during overnight for individual animals, using a computerised monitoring system; Pearson Technical Services, Debenham, Stowmarket, England). During the motor activity test, males were caged individually and females were caged with their offspring
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: No
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of dead and living pups at first Iitter check
Fetal examinations:
no data

OFFSPRING
Mortality/Viability: The numbers of live and dead pups at the First Lilter Check (=check at Day 1 of lactation) and daily thereafter were determined. lf possible, defects or cause of death were evaluated.
Clinical signs: At least once daily, detailed clinical observations (including abnormal behaviour) were made in all animals.
Body weights: Live pups were weighed during lactation on Days 1 and 4.
Sex: was determined for all pups on Days 1 and 4 of lactation (by assessment of the ano-genital distance).
Statistics:
- If the variables could be assumed to follow a normal distribution, the Dunnett-test (Dunnett, 1955) (many-to-one t-test) based on a pooled variance estimate was applied for the comparison of the treated groups and the control groups for each sex.
- The Steel-test (Miller, 1981) (many-to-one rank test) was applied if the data could not be assumed to follow a normal distribution.
- The Fisher Exact-test (Fisher, 1950) was applied to frequency data.
- No statistical analysis was performed on histopathology findings
All tests were two-sided and in all cases p < 0.05 was accepted as the lowest level of significance.
Historical control data:
- Results were compared with historical control data.
- No detailed data available in study

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:no effects

Details on maternal toxic effects:
This study is described in full detail under chapter 7.5.1 “Repeated dose toxicity: oral” and 7.8.1 "Toxicity to reproduction".

Effect levels (maternal animals)

open allclose all
Dose descriptor:
NOEL
Effect level:
150 mg/kg bw/day (nominal)
Basis for effect level:
other: developmental toxicity
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day (nominal)
Basis for effect level:
other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
This study is described in full detail under chapter 7.5.1 “Repeated dose toxicity: oral” and 7.8.1 "Toxicity to reproduction".

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
In conclusion, treatment with Isostearic acid, esters with methyl α-D-glucoside by oral gavage in male and female Wistar Han rats at dose levels of 0, 50, 150 and 1000 mg/kg bw/d revealed parental toxicity at 1000 mg/kg bw/day. No reproduction, breeding and developmental toxicity was observed for treatment up to 1000 mg/kg bw/day.
Executive summary:

In a Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test according OECD 422 test substance Isostearic acid, esters with methyl α-D-glucoside (100% UVCB substance (80% Methyl Glucoside Isostearate Esters (mainly Di-), 16% Isostearic Acid, 4% Methyl Glucoside)) in 1% aqueous carboxymethyl cellulose was administered to 10 male and 10 female Wistar Han rats/dose group by daily oral gavage at dose levels of 0, 50, 150, and 1000 mg/kg bw/day. The males were exposed for 2 weeks prior to mating, during mating, and up to termination (for 30 days). The females were exposed for 2 weeks prior to mating, during mating, post-coitum, and at least 4 days of lactation (for 42 to 44 days). 10 litters per dose group were delivered.

At 1000 mg/kg bw/day statistically significantly reduced haemoglobin, cholesterol and total protein levels (males), and elevated white blood cell counts (determined for only two females) plus alkaline phosphatase levels (males) were found. Increased liver weights (absolute and relative) were noted for high dose males and females.

No treatment-related changes were noted in any of the remaining parameters investigated in this study (i.e. reproduction, breeding, pup development, and of the adults: mortality, clinical appearance, functional observations, body weight, food consumption, macroscopic and microscopic examination,).

The parental NOEL is 150 mg/kg bw/day, based on the findings noted at 1000 mg/kg bw/day

The parental NOAEL is >= 1000 mg/kg bw/day based on the findings noted at 1000 mg/kg bw/day which were not considered adverse and were without any corroborative findings like histopathological changes.

The reproduction, breeding and developmental NOAELis >= 1000 mg/kg bw/d.

This Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test in the rat is acceptable and satisfies the guideline requirements of OECD TG 422.

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