Sucrose and glycerol, reaction products with C12-18, C18unsatd. fatty acids

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Link to relevant study records

Reference

Endpoint:

toxicity to reproduction

Remarks:

other: combined repeated dose and reproduction / developmental screening

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.

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)

Version / remarks:

March 1996

Deviations:

no

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

other: Wistar Han (Crl:WI(Han))

Sex:

male/female

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

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.

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).

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%.

Duration of treatment / exposure:

Offspring: not treated
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, during post-coitum, and during at least 4 days of lactation

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.

Details on study schedule:

- Parturition F0:
The females were allowed to Iitter normally. Day 1 of lactation was defined as the day when a litter was found completed (i.e. membranes, placentas cleaned up, nest built up and/or feeding of pups started). Females that were littering were left undisturbed.
- Lactation F0:
Examination of maternal care revealed no deficiencies (such as inadequate construction or cleaning of the nest, pups left scattered and cold, physical abuse of pups or apparently inadequate lactation or feeding).

Remarks:

Doses / Concentrations:
0, 50, 150 and 1000 mg/kg bw/day
Basis:
actual ingested

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Details on study design:

This study is described in full detail under chapter 7.5.1 "Repeated dose toxicity: oral".

Positive control:

no positive control group

Parental animals: Observations and examinations:

This study is described in full detail under chapter 7.5.1 "Repeated dose toxicity: oral".

Oestrous cyclicity (parental animals):

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
- Corpora lutea

Sperm parameters (parental animals):

All recorded microscopic findings were within the range of background pathology encountered in Wistar rats of this age in this type of study and occurred at similar incidences and severity in both control and treated rats. The spermatogenic staging profiles were normal for all group 1 and group 4 males evaluated.
Parameters examined in [all/P/F1/F2] male parental generations:
testis weight, epididymis weight, prostate weight, seminal vesicles, spermatogenesis staging

Litter observations:

PARAMETERS EXAMINED
The following parameters were examined in offspring:
- Number and sex of pups: on day 1 and 4 of lactation (by assessment of the ano-genital distance)
- Stillbirths, live births, postnatal mortality, if possible defects or cause of death: at day 1 of lactation and daily thereafter
- Clinical signs (detailed clinical observations, including abnormal behaviour): at least once daily
- Body weights: live pups were weighed during lactation on Days 1 and 4.

PATHOLOGY OFFSPRING
- Pups were killed by decapitation on Day 5 of lactation or shortly thereafter.
- The stomach was examined For the presence of milk.
- Descriptions of all external abnormalities were recorded. If possible, detects or cause of death were evaluated. Any abnormal pup, organ or tissue was preserved in 10% buffered formalin, for possible further examination.

Postmortem examinations (parental animals):

GROSS PATHOLOGY: Yes
- Dose groups: the first 5 mated males per group and the first 5 females with live offspring per group
-- Macroscopic examination: Adrenal glands, Aorta, Brain (cerebellum, mid-brain, cortex), Caecum, Cervix, Clitoral gland, Colon, Coagulation gland, Duodenum, Epididymides,
(Eyes with optic nerve (if detectable) and Harderian gland), (Female mammary gland area), (Femur including joint), Heart, Ileum, Jejunum, Kidneys, (Larynx), (Lacrimal gland, exorbital), Liver, Lung, Ovaries, Pancreas, Peyer's patches (jejunum, ileum) if detectable, Pituitary gland, Preputial gland, Prostate gland, Rectum, (Salivary glands - mandibular, sublingual), Sciatic nerve, Seminal vesicles, (Skeletal muscle), (Skin), Spinal cord -cervical, midthoracic, lumbar, Spleen, Sternum with bone marrow, Stomach, Testes, Thymus, Thyroid includinq parathyroid (if detectable), (Tongue), Trachea, Urinary bladder, Uterus, Lymph nodes - mandibular, mesenteric, (Nasopharynx), Oesophagus, Vagina, all gross lesions

- From all remaining animals:
Cervix, Clitoral gland, Coagulation gland, Epididymides,Ovaries, Preputial gland, Prostate gland, Seminal vesicles, Testes, Uterus, Vagina, all gross lesions

- Tissues/organs mentioned in parentheses were not examined by the pathologist, since no signs of toxicity were noted at macroscopic examination.

-- Organ weights:
- From the first 5 mated males per group and the first 5 females with live offspring per group: Adrenal glands, Brain, Epididymides, Heart, Kidneys, Liver, Prostate (when fixed for at least 24 hours), Seminal vesicles, Spleen, Testes, Thymus

- From all remaining males:
Epididymides, Testes, Prostate (when fixed for at least 24 hours), Seminal vesicles


HISTOPATHOLOGY: Yes
- Histopathologic examination was performed on an extensive list of organs and tissues from five males and five females of groups 1 and 4 as well as gross lesions from all rats. Sections of testes from five group 1 and 4 rats were assessed for spermatogenesis staging.
- Adrenal glands, aorta, bone - sternum [and femur including joint]; bone marrow - sternal, brain, clitoral glands, epididymides, esophagus, [eyes with optic nerve and Harderian glands); heart, [identification marks], kidneys, [Iacrimal glands - exorbital], large intestine cecum, colon and rectum; [larynx), liver, lungs, Iymph nodes - mandibular and mesenteric; [female mammary gland area], [nasopharynx], ovaries, pancreas, pituitary gland, preputial, glands, prostate gland, [salivary glands - mandibular and sublingual]; sciatic nerve, seminal vesicles with coagulation glands, [skeletal muscle], [skin], small intestine - duodenum, jejunum and ileum with Peyer's patches: spinal cord - cervical, midthoracic and lumbar; spleen, stomach, testes, thymus, thyroid glands with parathyroid glands, [tongue], trachea, urinary bladder, uterus with uterine cervix, vagina and all organs or tissues with macroscopic abnormalities.
Following fixation, organs (except those listed in brackets) from the selected animals of groups 1 and 4 along with all organs or tissues with macroscopic abnormalities from all rats, were trimmed , processed and embedded in paraffin wax, precision cut and stained with hematoxylin and eosin.

Postmortem examinations (offspring):

Pups were killed by decapitation on Day 5 of lactation or shortly thereafter.

All offspring was sexed and externally examined. The stomach was examined for the presence of milk. Descriptions of all external abnormalities were recorded. If possible, defects or cause of death were evaluated. Any abnormal pup, organ or tissue was preserved in 10% buffered formalin, for possible further examination.
No treatment-related changes were noted for reproduction, breeding and pup development.

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.

Clinical signs:

no effects observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

no effects observed

Other effects:

effects observed, treatment-related

Reproductive function: oestrous cycle:

no effects observed

Reproductive function: sperm measures:

no effects observed

Reproductive performance:

effects observed, treatment-related

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
- Corpora lutea

REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS)
- Staging of Spermatogenesis

For further details please find under chapter: 7.5.1 "Repeated dose toxicity: oral".

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

other: see 'Remark'

Dose descriptor:

NOEL

Effect level:

150 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

other: see 'Remark'

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

other: - No treatment-related changes on reproduction, breeding and pup development).

Remarks on result:

other: Generation: reproduction, breeding, development (migrated information)

Clinical signs:

no effects observed

Mortality / viability:

no mortality observed

Body weight and weight changes:

no effects observed

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Histopathological findings:

not examined

VIABILITY (OFFSPRING)
- Viability index (=Number of alive pups before planned necropsy/Number of pups born alive): 0 mg/kg bw/d: : 100% , 50 mg/kg bw/d: : 100%, 150 mg/kg bw/d: : 99.2%, 1000 mg/kg bw/d: : 98.4% (significant at 5 %)
- Dead pups at first litter check: 0 mg/kg bw/d: 3 males, 2 females; 50 mg/kg bw/d: 1 male, 1 female, 150 mg/kg bw/d: 0; 1000 mg/kg bw/d: 1 female
- Dead pups post natal: at 0, 50 mg/kg bw/d: 0, at 150 mg/kg bw/d: 1 male, at 1000 mg/kg bw/d: 1 male, 1 female from one litter
- 10 litters/dose group

CLINICAL SIGNS (OFFSPRING)
- small size, bluish colour, blue spot on the neck and eye, scabbing of the right cheek, pale appearance and insufficient milk in the stomach. No relationship with treatment was established for these observations and they were considered to be of no toxicoiogical significance.

BODY WEIGHT (OFFSPRING)
Pup (mean) body weights were in the same range for the control and treated groups.

GROSS PATHOLOGY (OFFSPRING)
- Findings consisted of autolysis of pups found dead at the first Iitter check, scabbing of the right cheek, and insufficient milk in the stomach. No relationship with treatment was established for these observations and they were considered to be of no toxicoiogical significance.

Reproductive effects observed:

not specified

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/day 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 Isostearic acid, esters with methyl α-D-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, during 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 NOAEL is >= 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.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

Effects on fertility are assessed on the basis of two guideline studies. The key study is a combined repeated dose and reproduction / developmental screening study (OECD 422) performed on a supporting substance (structural analogue). The supporting study is a 90 day repeated dose oral toxicity study (OECD 408) performed on a source substance and constituent.

Both studies reported are high quality guideline compliant studies for which the Klimisch score has been modified from RL1 to RL2 due to the fact that the studies were performed on read-across substances.

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

Key study on the read-across substance Isostearic acid, esters with methyl α-D-glucoside (Evonik, 2009)

The study was performed according to OECD guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test) with Isostearic acid, esters with methyl α-D-glucoside by daily exposure in the rat.

The repeated dose part of this study has been used as reference for subacute repeated dose toxicity.

The reproduction/developmental toxicity screening part of this study has been used as reference for reproductive toxicity (fertility) and for developmental toxicity.

Based on the results of a 14-day range finding study, the dose levels for the subacute treatment were selected to be 0, 50, 150 and 1000 mg/kg bw/day. Isostearic acid, esters with methyl α-D-glucoside in 1% aqueous carboxymethyl cellulose was administered daily by oral gavage to groups of 10 male and 10 female Wistar Han rats 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, during post-coitum, and at least 4 days of lactation (for 42 to 44 days). 10 litters per dose group were delivered. The in-life phase was concluded on day 5 of lactation.

The stability of the test substance in the vehicle was determined during the project. Chemical analyses of formulations were conducted once during the study to assess accuracy and homogeneity. The following parameters were evaluated: clinical signs daily; functional observation tests in week 4; body weight and food consumption weekly; clinical pathology and macroscopy at termination; organ weights and histopathology on a selection of tissues.

Further to the exams performed on parental animals, the offspring was examined for viability, clinical signs, body weight, sexual maturation, organ weights, gross pathology and histopathology.

 

Results

No mortality occurred during the study period.

Parental animals:

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.

Parental animals and offspring:

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

There was no evidence of reproduction/developmental toxicity in response to treatment with the test substance.

 

Conclusion

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 NOAEL is >= 1000 mg/kg bw/day, based on the absence of treatment-related changes on reproduction, breeding and pup development.

 

 

Supporting study on the source substance and constituent Mono-Di-Glycerides (Irwin, 1992)

This study has been used as reference for subchronic repeated dose toxicity.

Itis a 90 day oral (feeding) study performed in rats and mice with a Mono-Di-Glyceride, Castor Oil, a read-across substance (source substance and constituent). The study is equivalent to the OECD guideline 408 (Repeated Dose 90-day Oral Toxicity Study in Rodents), but no ophthalmoscopy and no neurologic exams were performed.

Castor oil is a natural oil derived from the seeds of the castor bean, Ricinus communis. It is comprised largely of triglycerides with a high ricinolin content. Toxicity studies with castor oil were performed by incorporating the material at concentrations as high as 10% in diets given to F344/N rats and B6C3F1 mice of both sexes (10 males and 10 females per dose group) for 13 weeks.

 

Conclusion

A NOAEL of 5000 mg/kg bw/day for rats and a NOAEL of 15000 mg/kg bw/day for mice could be identified based on the absence of any treatment-related changes for parental fertility parameters.

 

 

General evaluation of toxicity to reproduction - fertility

In a reproduction/developmental toxicity screening study on Isostearic acid, esters with methyl α-D-glucoside, a read-across substance (structural analogue), the reproduction, breeding and developmental NOAEL is >= 1000mg/kg bw/day, based on the absence of treatment-related changes on reproduction, breeding and pup development.

The toxicological characteristics of this read-across substance have been shown to be comparable to the properties of the registration substance, as referred for the endpoints acute oral toxicity, skin and eye irritation, skin sensitisation and genetic toxicity (Ames Test).

In a subchronic study on a Mono-Di-Glyceride, Castor Oil, a read-across substance (source substance and constituent), the NOAELs observed in this study were 5000 mg/kg bw/day for rats and 15000 mg/kg bw/day for mice, based on the absence of any treatment-related changes for parental fertility parameters.

The information presented is considered complete and relevant with regard to the possible routes of exposure and for the assessment of risk in humans.

The results of the studies for the endpoint toxicity to reproduction, i.e. the complete absence of adverse effects and reasons for concern, cover the information requirements of the REACH Regulation Annex IX for reproductive toxicity / fertility and can be considered reliable and consistent.

The results obtained in all studies clearly show that no adverse effects have been observed at dose levels >= 1000 mg/kg bw/day.

Extrapolating the results from the read-across substances to the registration substance, it is concluded thatSucroglyceride C12-18, C18unsatd. has a reproduction, breeding and developmental NOAEL>= 1000 mg/kg bw/day and that the substancedoes not require classification and labelling for damage to health by toxicity to reproduction (fertility) or by developmental toxicity.

Short description of key information:
In a Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test according to OECD 422, performed on a read-across test substance (structural analugue), no reproduction, breeding and developmental toxicity was observed for treatment up to 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 NOAEL is >= 1000 mg/kg bw/day.

In a 90 day oral dietary toxicity study the read across test substance (source substance and constituent) was administered to rats and mice, to groups of 10 animals/sex/dose at concentrations of up to 10 % in the diet. A NOAEL of 5000 mg/kg bw/day for rats and a NOAEL of 15000 mg/kg bw/day for mice could be identified based on parental fertility parameters.

Justification for selection of Effect on fertility via oral route:
The key study chosen, performed on a read-across substance (structural analogue) is a subacute study. Only this one reproduction toxicity study is available. The results show the absence of treatment-related changes for reproduction, breeding and pup development.

Furthermore, a supporting subchronic (90 day repeated dose) study is presented, performed on a source substance / constituent. During this study, parental fertility parameters have been assessed and no adverse effects have been identified.

The results presented in the subacute key and in the subchronic supporting study provide evidence for the absence of adverse effects on reproductive organs and tissues. Hence, according to the REACH regulation, Annex IX 8.7.3., column 1, the performance of a two generation reproductive toxicity study is not considered necessary.

Justification for selection of Effect on fertility via inhalation route:
Exposure of humans via inhalation is unlikely taking into account the possibility of exposure to aerosols, particles or droplets of an inhalable size. Sucroglyceride C12-18, C18unsatd. is a waxy solid paste. Taking this consistence into account the generating of inhalable particles such as dust or aerosols is not significant due to the specific operational conditions.

Justification for selection of Effect on fertility via dermal route:
Systemic exposure of humans by the dermal route is unlikely taking into account that a significant rate of absorption through the skin cannot be expected.

Effects on developmental toxicity

Description of key information

In a Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test according to OECD 422, performed on a read-across test substance (structural analugue), no reproduction, breeding and developmental toxicity was observed for treatment up to 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 NOAEL is >= 1000 mg/kg bw/day. 

Link to relevant study records

Reference

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.

Reason / purpose for cross-reference:

reference to same study

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

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

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

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.

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

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".

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

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".

Abnormalities:

not specified

Developmental effects observed:

not specified

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.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

Effects on developmental toxicity are assessed on the basis of one key study. It is a guideline study, combined repeated dose and reproduction / developmental screening, performed on a supporting substance (structural analogue).

The toxicological characteristics of the read-across substance examined in the key study have been shown to be comparable to the properties of the registration substance, as referred for the endpoints acute oral toxicity, skin and eye irritation, skin sensitisation and genetic toxicity (Ames Test).

The study presented for this endpoint covers the Annex IX testing requirements of the REACH Regulation for pre-natal developmental toxicity and can be considered reliable and consistent. It is a high quality guideline compliant studies for which the Klimisch score has been modified from RL1 to RL2 due to the fact that the study was performed on read-across substances.

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Quality of whole database:

Effects on developmental toxicity are assessed on the basis of one guideline studies. The key study is a combined repeated dose and reproduction / developmental screening study performed on a supporting substance (structural analogue).

The toxicological characteristics of the read-across substance examined in the key study have been shown to be comparable to the properties of the registration substance, as referred for the endpoints acute oral toxicity, skin and eye irritation and skin sensitisation.

The study presented for this endpoint covers the Annex IX testing requirements of the REACH Regulation for pre-natal developmental toxicity and can be considered reliable and consistent. It is a high quality guideline compliant studies for which the Klimisch score has been modified from RL1 to RL2 due to the fact that the study was performed on read-across substances.

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

Key study on the read-across substance Isostearic acid, esters with methyl α-D-glucoside (Evonik, 2009)

The study was performed according to OECD guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test) with Isostearic acid, esters with methyl α-D-glucoside by daily exposure in the rat.

The repeated dose part of this study has been used as reference for subacute repeated dose toxicity.

The reproduction/developmental toxicity screening part of this study has been used as reference for reproductive toxicity (fertility) and for developmental toxicity.

Based on the results of a 14-day range finding study, the dose levels for the subacute treatment were selected to be 0, 50, 150 and 1000 mg/kg bw/day. Isostearic acid, esters with methyl α-D-glucoside in 1% aqueous carboxymethyl cellulose was administered daily by oral gavage to groups of 10 male and 10 female Wistar Han rats 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, during post-coitum, and at least 4 days of lactation (for 42 to 44 days). 10 litters per dose group were delivered. The in-life phase was concluded on day 5 of lactation.

The stability of the test substance in the vehicle was determined during the project. Chemical analyses of formulations were conducted once during the study to assess accuracy and homogeneity. The following parameters were evaluated: clinical signs daily; functional observation tests in week 4; body weight and food consumption weekly; clinical pathology and macroscopy at termination; organ weights and histopathology on a selection of tissues.

Further to the exams performed on parental animals, the offspring was examined for viability, clinical signs, body weight, sexual maturation, organ weights, gross pathology and histopathology.

 

Results

No mortality occurred during the study period.

Parental animals:

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.

Parental animals and offspring:

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

There was no evidence of reproduction/developmental toxicity in response to treatment with the test substance.

 

Conclusion

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 NOAEL is >= 1000mg/kg bw/day, based on the absence of treatment-related changes on reproduction, breeding and pup development.

 

General evaluation of developmental toxicity / teratogenicity

In a reproduction/developmental toxicity screening study on Isostearic acid, esters with methyl α-D-glucoside, a read-across substance (structural analogue), the reproduction, breeding and developmental NOAEL is >= 1000mg/kg bw/day, based on the absence of treatment-related changes on reproduction, breeding and pup development.

The toxicological characteristics of this read-across substance have been shown to be comparable to the properties of the registration substance, as referred for the endpoints acute oral toxicity, skin and eye irritation, skin sensitisation and genetic toxicity (Ames Test).

The information presented is considered complete and relevant with regard to the possible routes of exposure and for the assessment of risk in humans.

The results of the studies for the endpoint toxicity to reproduction, i.e. the complete absence of adverse effects and reasons for concern, cover the information requirements of the REACH Regulation Annex IX for reproductive toxicity / fertility and can be considered reliable and consistent.

The results obtained in all studies clearly show that no adverse effects have been observed at dose levels >= 1000 mg/kg bw/day.

Extrapolating the results from the read-across substances to the registration substance, it is concluded thatSucroglyceride C12-18, C18unsatd. has a reproduction, breeding and developmental NOAEL >= 1000 mg/kg bw/day and that the substancedoes not require classification and labelling for damage to health by toxicity to reproduction (fertility) or by developmental toxicity.

Justification for selection of Effect on developmental toxicity: via oral route:
The key study chosen, performed on a read-across substance (structural analogue) is a subacute study. Only this one reproduction toxicity study is available.

The study shows the absence of treatment-related changes for reproduction, breeding and pup development. There is no concern for pre-natal developmental toxicity. Hence the information requirements of the REACH regulation, Annex IX 8.7.2., column 1 are satisfied and further testing for pre-natal developmental toxicity is not considered necessary.

Justification for selection of Effect on developmental toxicity: via inhalation route:
Exposure of humans via inhalation is unlikely taking into account the possibility of exposure to aerosols, particles or droplets of an inhalable size. Sucroglyceride C12-18, C18unsatd. is a waxy solid paste. Taking this consistence into account the generation of inhalable particles such as dust or aerosols is not significant due to the specific operational conditions.

Justification for selection of Effect on developmental toxicity: via dermal route:
Systemic exposure of humans by the dermal route is unlikely taking into account that a significant rate of absorption through the skin cannot be expected.

Justification for classification or non-classification

In a read-across study, for rats, performed on Isostearic acid, esters with methyl α-D-glucoside, a strucural analogue, a parental NOAEL >= 1000 mg/kg bw/day and a reproduction, breeding and developmental NOAEL >= 1000 mg/kg bw/day were observed. These results have been considered for toxicity to reproduction (fertility) and for developmental toxicity.

The read-across data can be extrapolated to the registration substance Sucroglyceride C12 -18, C18unsatd., especially since the toxicological characteristics of the read-across substance of the subacute study have been shown to be comparable to the properties of the registration substance, as referred for the endpoints acute oral toxicity, skin and eye irritation, skin sensitisation and genetic toxicity (Ames Test). 

Furthermore, in a 90 day oral study performed on Castor Oil, a source substance and constituent, NOAELs were reported to be 5000 mg/kg bw/day for rats and 15000 mg/kg bw/day for mice, based on the absence of any treatment-related changes for parental fertility parameters.

 

Extrapolating the read-across data to the registration substance, regarding a potential damage to reproductive functions and the development of offspring, according to the CLP Regulation (EC) No 1272/2008 and according to DSD (67/548/EEC), there is no need for classification of Sucroglyceride C12 -18, C18unsatd. and no labelling is required.