2,2-bis(hydroxymethyl)propane-1,3-diyl didocosanoate

Endpoint summary

Description of key information

Oral: OECD 408, rat, NOAEL ≥ 1000 mg/kg bw/day

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: oral

Type of information:

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

Adequacy of study:

key study

Study period:

17 Nov 1997 - 16 Mar 1998

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

GLP - Guideline study, teste with the source substance Isononanoic acid, mixed esters with 2-methylbutanoic acid, 3-methylbutanoic acid, pentaerythritol and valeric acid. According to the ECHA guidance document "Practical guide 6: How to report read-across and categories' ECHA, 2012), the reliability was changed from RL1 to RL2 to reflect the fact that this study was conducted on a read-across substance.

Qualifier:

according to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

ToxLabs Prüflabor GmbH, Greppin, Germany

Limit test:

no

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Wiga GmbH
- Age at study initiation: 32-38 d
- Weight at study initiation: 148.5 g (mean value males), 136.7 g (mean value females)
- Housing: one or two animals in cages (Makrolon Type 3)
- Diet: Altromin 1326, ad libitum
- Water: tap water, ad libitum
- Acclimation period: 6-8 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-23
- Humidity (%): 30-60
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: gavage

Vehicle:

other: distilled water containing 1% Tween 80

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 1%
- Lot/batch no. (if required): S23350 739

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Duplicate 2 mL samples of each formulation were taken and stored in the frozen state until measurement

Duration of treatment / exposure:

90 d

Frequency of treatment:

once daily, 7 days/week

Dose / conc.:

100 mg/kg bw/day (actual dose received)

Dose / conc.:

300 mg/kg bw/day (actual dose received)

Dose / conc.:

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

No. of animals per sex per dose:

10 (control, test and satellite groups)

Control animals:

yes, concurrent vehicle

Details on study design:

- Rationale for selecting satellite groups: 10 animals each from the high dose and the vehicle group were used to investigate reversibility of possible effects
- Post-exposure recovery period in satellite groups: 28 d

Positive control:

No

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes, autonomic activity, presence of clonic or tonic movements, stereotypies, bizarre behavior
- Time schedule: Daily

DETAILED CLINICAL OBSERVATIONS: Yes, changes in skin, fur, eyes, mucous membranes, gait, posture: response to handling; occurrence of secretions and excretions
- Time schedule: Daily

BODY WEIGHT: Yes
- Time schedule for examinations: Weekly from the start to the end of the study

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: prior to the administration and at the end of the study
- Dose groups that were examined: All (surviving) animals

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Just prior to killing at the end of the study
- Anesthetic used for blood collection: Yes (Ether)
- Animals fasted: Yes, over night
- How many animals: All animals
- Parameters examined: erythrocyte count, hemoglobin concentration, packed cell volume, platelet count, total leukocyte count, leukocyte differential count, prothrombine time, fibrinogen concentration

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: Just prior to killing at the end of the study (including the satellite groups)
- Animals fasted: Yes
- How many animals: all animals
- Parameters examined: alkaline aminotransferase, alkaline phosphatase, aspartate aminotransferase, bilirubin, blood urea nitrogen, calcium, creatinine, fasting glucose, phosphorus, total cholesterol, total protein, albumin, chloride, potassium, sodium

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: Prior to administration, at monthly intervals and in the last week of dosing and in the last week of the recovery period.
- Dose groups that were examined: all
- Battery of functions tested: sensory activity (auditory, visual and proprioceptive stimuli) / grip strength / motor activity

Sacrifice and pathology:

GROSS PATHOLOGY: cranial, thoracic and abdominal cavities were opened and examined macroscopically
HISTOPATHOLOGY: adrenals, aorta, brain (3 sections), epididymides, eye, femur, heart, kidney, liver, lungs (incl. mainstem bronchi), mesenteric lymph node, muscle incl. sciatic nerve, oesophagus, ovaries, pancreas, pituitary, prostate, seminal vesicle, skin incl. mammary glands, small and large intestine (including peyer´s patches), spinal chord (3 levels), spleen, sternum with bone marrow, stomach, submandibular lymph node, testes, thymus, thyroid (incl. parathyroids), trachea, urinary bladder, uterus

Other examinations:

Organ weights of adrenals, brain, epididymides, testes, heart, kidneys, liver, ovaries, spleen, testes and thymus

Statistics:

Body weights, food consumption: Welch t-test

Haematology, coagulation, clinical biochemistry and absolute and relative organ weights: Dunnett´s test

Differential leukocyte count: Mean, range and standard deviation

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

no effects observed

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

high dose group (female): fibrinogen concentration of plasma and creatinine content increased, high dose group (male/female): alkaline phosphatase increased, middle and high dose groups (male/female): sreum urea nitrogen increased, all non-adverse

Urinalysis findings:

not examined

Behaviour (functional findings):

no effects observed

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

increased kidney weights for high dose males and females, non-adverse

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

intracellular fat and low-grade fatty degenerations of hepatocytes in all male animals, female control, middle dose and high dose groups, non-adverse

Histopathological findings: neoplastic:

no effects observed

Details on results:

CLINICAL SIGNS AND MORTALITY
Two animals died shortly after administration due to incorrect gavage shown by lungs filled with blood. None of the animals showed any alterations of their general state of well-being and behaviour at any observation period (few observations were made substance independent and for a short period of time).

BODY WEIGHT AND WEIGHT GAIN
Not affected by the test compound.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
Not dose-dependently influenced.

OPHTHALMOSCOPIC EXAMINATION
No alterations.

HAEMATOLOGY
Not influenced.

CLINICAL CHEMISTRY
The fibrinogen concentration of the plasma was increased in the female animals of the high dose group, this was no longer apparent at the end of the treatment-free period.
The activity of alkaline phosphatase of the serum significantly increased in the high dose group, males and femals. This indicates damage to liver cells and/or an increased function rate. This finding was no longer apparent at the end of the treatment-free period.
The serum urea nitrogen was significantly increased in the middle and high dose group of the males and in the high dose group of female animals. The creatinine content was significantly increased in all male and in the high dose group of the female animals. The phosphorus content was significantly increased dose-dependently in all female animals and the sodium content was dose-dependently decreased in the male animals, significantly in the animals of the middle and high dose groups. These effects were no longer apparent at the end of the treatment-free period.

NEUROBEHAVIOUR
No changes in grip strength, motor activity and sensory response.

ORGAN WEIGHTS
Absolute and relative kidney weights were increased in all male animals in the high dose group which was still present after the recovery period. Absolute and relative liver weights were increased in both sexes but this was no longer apparent after the recovery period in females. Other significant differences seem to be incidental.

GROSS PATHOLOGY
No substance-dependent changes.

HISTOPATHOLOGY: NON-NEOPLASTIC
Intracellular lipid droplets in hepatocytes of the female animals in the high and mid dose group (5-90% of the observed area) with cell lesions were clearly caused dose-dependently by the test article. There was no special localization of the changes of hepatocytes in the liver lobules. In most cases only low grade intracellular lipopexia occurred in the male animals.
Stomach: Oesophagal part and cardia with multilayered squamous epithelium, leukocyte infiltration in the submucosa and fibrous repair, fibrotic regions in the submucosa of the glandular stomach
Lungs: Atelectactic and emphysemic areas
Thymus: Partial substitution of the parenchyma by fibrinogenesis
Skin: Benign fibrous proliferation
Sciatic nerve: Thickening of the perineurium and thickening of the adventitia of the vessels

Dose descriptor:

NOAEL

Effect level:

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

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no adverse effects observed

Dose descriptor:

NOEL

Effect level:

300 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

clinical biochemistry

organ weights and organ / body weight ratios

Critical effects observed:

not specified

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Quality of whole database:

The available information comprises adequate, reliable (Klimisch score 2 due to read-across) and consistent studies from a reference substance with similar structure and intrinsic properties. Read-across is justified based on common precursors and breakdown products of hydrolysis and consistent trends in environmental fate, ecotoxicological and toxicological profile (refer to the endpoint discussion for further details).
The selected study is thus sufficient to fulfil the standard information requirements set out in Annex VIII-IX, 8.6, in accordance with Annex XI, 1.5, of Regulation (EC) No 1907/2006.

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Read across justification

There are no data on the repeated dose toxicity of 2,2-bis(hydroxymethyl)propane-1,3-diyl didocosanoate (CAS 68258-72-0). The assessment was therefore based on studies conducted with analogue substances as part of a read across approach, which is in accordance with Regulation (EC) No. 1907/2006, Annex XI, 1.5. For each specific endpoint the source substance(s) structurally closest to the target substance is/are chosen for read-across, with due regard to the requirements of adequacy and reliability of the available data. Structural similarities and similarities in properties and/or activities of the source and target substance are the basis of read-across. A detailed justification for the analogue read-across approach is provided in the technical dossier (see IUCLID Section 13).

Repeated dose toxicity, oral, subchronic

CAS 146289-36-3

A 90-day oral repeated dose toxicity study was performed according to OECD guideline 408 and under GLP conditions, with pentaerythritol ester of pentanoic acids and isononanoic acid (Müller, 1998). 10 Wistar rats/sex/dose were administered 100, 300 and 1000 mg/kg bw/day by gavage, for 90 consecutive days. Satellite control and high dose groups of 10 animals/sex/dose were observed for 28 days after test substance administration ended. While no toxicologically relevant mortality was observed, two animals died shortly after administration due to an incorrect gavage procedure. No clinical signs were observed during the study period. No significant differences in body weight, body weight gain and food consumption between the control group and treatment groups were noted. The ophthalmoscopic examination did not show any treatment-related changes in any group. The hematology results were comparable in the control and treatment groups. No significant effects were observed in the neurobehavioural assessment. The alkaline phosphatase level was significantly increased in the high dose group, in males and females. This increase in a liver enzyme may be caused by an increase in the liver metabolism due to the test substance. Furthermore, the absolute and relative liver weights were increased in both sexes in the main group. However, no other effects on hepatic functions were seen on clinical chemistry parameters or in the histopathological results, and the effect on AST levels (males and females) and liver weight (males) were reversible. Therefore the effect is considered to be adaptive. The serum urea nitrogen was significantly increased in the mid- and high dose group of the males and in the high dose group of female animals. The creatinine content was significantly increased in all male and in the high dose group of the female animals. These effects were no longer observed at the end of the treatment-free period. Due to the reversibility and due to the absence of other relevant findings, these changes are not considered to be toxicologically relevant. The absolute and relative kidney weights were increased in all male animals in the high dose main group and satellite group. This is most likely correlated to the formation of hyaline droplets, which is a well-known effect specific to the male rat and as such considered to have no relevance to humans. Intracellular lipid droplets in hepatocytes of females in the high and mid dose group (5-90% of the observed area) with cell lesions were apparently caused dose-dependently by the test substance. There was no special localization of the changes of hepatocytes in the liver lobules. In most cases only low grade intracellular lipopexia occurred in the male animals. The effect is treatment-related but not toxicologically relevant. The macroscopic inspection at autopsy and subsequent histopathological examination of remaining organs and tissues did not reveal any treatment-related changes. The NOAEL for systemic toxicity was considered to be ≥ 1000 mg/kg bw/day.

CAS 68424-31-7

Furthermore, a 28 day study with Fatty acids, C5-10, esters with pentraerythritol (CAS 68424-31-7) was conducted according to OECD Guideline 407 and under GLP conditions (Brammer, 1993). The test substance was administered in concentrations of 1000 ppm, 5000 ppm and 12500 ppm (corresponding to 112, 562 and 1450 mg/kg bw/day for male and 119, 586 and 1613 mg/kg bw/day for female rats) to 5 Alpk:APfSD rats per sex and dose for 28 consecutive days. Control animals (5 per sex and dose) received the plain diet. There were no toxicologically significant effects on body weight, food consumption and clinical condition and mortality up to and including the highest dose level. Changes in clinical chemistry and red cell-related parameters were observed in male rats at 12500 ppm, but these were minor and considered not to be of toxicological significance. A minimal hepatocyte hypertrophy present in males of the 12500 ppm group was observed and considered to be evidence of an adaptive response. Microscopic examination of the kidneys from male animals from all dose groups revealed an increase in hyaline droplet formation (the main constituent of which is alpha-2µ-globulin) and tubular basophilia. This phenomenon is widely accepted to be specific to the male rat and as such is considered to have no relevance to man. A NOAEL of 1450 and 1613 mg/kg/d could be identified for male and female rats, respectively.

Overall conclusion for repeated dose toxicity

The data for the read-across analogue substances showed that no effects were observed up to and including the recommended limit values. Therefore, as the available data did not identify any hazard for repeated dose toxicity, 2,2-bis(hydroxymethyl)propane-1,3-diyl didocosanoate is not expected to be hazardous following repeated exposure.

Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Hazard assessment is conducted by means of read-across from structural analogues. The selected study is the most adequate and reliable study based on the identified similarities in structure and intrinsic properties between the source and target substance and overall assessment of quality, duration and dose (refer to the endpoint discussion for further details).

Justification for classification or non-classification

According to Article 13 of Regulation (EC) No. 1907/2006 "General Requirements for Generation of Information on Intrinsic Properties of substances", information on intrinsic properties of substances may be generated by means other than tests e.g. from information from structurally related substances (grouping or read-across), provided that conditions set out in Annex XI are met. Annex XI, "General rules for adaptation of this standard testing regime set out in Annexes VII to X” states that “substances whose physicochemical, toxicological and ecotoxicological properties are likely to be similar or follow a regular pattern as a result of structural similarity may be considered as a group, or ‘category’ of substances. This avoids the need to test every substance for every endpoint". Since the analogue concept is applied to 2,2-bis(hydroxymethyl)propane-1,3-diyl didocosanoate (CAS 68258-72-0), data will be generated from data for reference source substance(s) to avoid unnecessary animal testing. Additionally, once the analogue read-across concept is applied, substances will be classified and labelled on this basis.

Therefore, based on the analogue read-across approach, the available data on repeated dose toxicity do not meet the classification criteria according to Regulation (EC) 1272/2008, and are therefore conclusive but not sufficient for classification.