Reaction mass of glycerol 1,3-di(acetate) and glycerol acetate and triacetin

Endpoint summary

Description of key information

Repeated dose toxicity: Oral NOAEL (rat, m/f): 1000 mg/kg bw/day (OECD 422, GLP)

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

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

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

GLP-Guideline study, tested with the source substance Triacetin (CAS No 102-76-1). According to 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)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

other: Crj: CD (SD) IGS

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 9 weeks
- Weight at study initiation: 317-375 g (males) and  203-240 g (females)

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-25
- Humidity (%): 31-62
- Air changes (per hr): 12
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: gavage

Vehicle:

other: 3% gum arabicum in purified water

Details on oral exposure:

Dosing volume: 5 mL/kg bw

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

44 days from 14 days prior to mating (males)
41-48 days from 14 days before mating to day 3 postpartum (females)

Frequency of treatment:

once daily, 7 days/week

Remarks:

Doses / Concentrations:
40, 200, 1000 mg/kg bw/day
Basis:
actual ingested

No. of animals per sex per dose:

12

Control animals:

yes, concurrent vehicle

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
- Time schedule for examinations: on Day 0, 3, 7 and 14 of administration and once a week thereafter. For pregnant females, body weight was determined on Day 0, 7, 14 and 20 of gestation and on Day 0 and 4 of lactation.

FOOD CONSUMPTION: Yes
- Food consumption was determined on the same day when body weight was measured for 24 h.

HAEMATOLOGY: Yes
- Time schedule for examinations: at time of necropsy after 44 days of chemical exposure (males).
- Parameters checked: red blood cell count, haemoglobin, haematrocrit, mean corpuscular volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, reticulocyte ratio, platelets, white blood cell count, lymphocyte, neutrophilic segmented, neutrophilic band, eosinophil-, basophil-, monocyte-value.

CLINICAL CHEMISTRY: Yes
- Time schedule for examinations: at time of necropsy after 44 days of chemical  exposure (males).
- Parameters checked: aspartate aminotransferase, alanine aminotrasferase, gamma glutamyltransferase, nitrophenylphosphate, total bilirubin, urea nitrogen, creatinine, glucose, total cholesterol, triglycerides, total protein, albumin, A/G ratio, calcium, inorganic phophorus, Na, K, Cl.

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
Organs examined at necropsy: organ weight for both sexes, brain, pituitary gland, thyroid gland, heart, liver, kidney, spleen, adrenal, thymus and in addition for males, testes and epididymis.

HISTOPATHOLOGY: Yes
All animals in control and in the high dose group and unfertilized animals in other groups: brain, spinal cord, pituitary gland, eyeball, thyroid 
gland (including parathyroid gland), thymus, heart, trachea, lung, liver, kidney, adrenal, spleen, stomach, small  intestine, large intestine, pancreas, 
urinary bladder, bone marrow, sciatic nerve, lymph node, testes, epididymis, prostate, seminal vesicle,  ovary, uterus, vagina, mammary gland and any organs, which might be expected to have histopathological changes and thymus and lung of dead animals.

Statistics:

Regarding quantitative data (body weight, gain of body weight, food consumption, organ weight, haematology, clinical chemistry, number of corpora lutea, number of implantation sites and total number of offspring), Bartlett test was used. In case of equal variance and unequal variance, ANOVA and Kruskal-Wallis test was applied, respectively. If there was a significant difference, Dunnett test or Dunnett multiple-comparison was used.
For day of conceiving, number of estrus, gestation length, implantation index, delivery index, viability index at Day 0 and Day 4, Bartlett test and Kruskal-Wallis test was used. If a significant difference was found, Dunnett multi-comparison test was applied.
For histopathology findings, the Chi-square test was used. If a significant difference was observed, Chi-square of Armitage was used between control and administration group. Regarding copulation index, fertility index, gestation index and sex ratio of offspring was tested with Fisher’s exact test.

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:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Details on results:

CLINICAL SIGNS AND MORTALITY
No clinical signs of toxicity observed. One male at 1000 mg/kg bw/day was dead 32 days after the administration started.

BODY WEIGHT AND WEIGHT GAIN
No effects were observed.

FOOD CONSUMPTION AND COMPOUND INTAKE
No effects were observed.

HAEMATOLOGY
In males a decrease in differential leukocyte count (%) in neutrophils (band) at 40 (p < 0.05) and 1000 mg/kg bw/day (p < 0.01) was observed, which was within physiological changes.

CLINICAL CHEMISTRY
In males a decrease in creatinine at 40 (p < 0.01) and 1000 mg/kg bw/day (p < 0.01) was observed,  which was within physiological changes. An increase in inorganic phosphorus at 200 mg/kg bw/day (p < 0.05) was apparent, but with no dose-related changes.

ORGAN WEIGHTS
No changes were observed.

GROSS PATHOLOGY
No changes were observed.

HISTOPATHOLOGY: NON-NEOPLASTIC
No changes were observed.

Dose descriptor:

NOAEL

Effect level:

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

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: overall effects

Critical effects observed:

not specified

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

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

Justification for grouping of substances and read-across

There are no data available on the repated dose toxicity of "Reaction mixture of glycerol-1,3-di(acetate), glycerol acetate and triacetin".

In order to fulfil the standard information requirements set out in Annex VIII, 8.6, in accordance with Annex XI, 1.5, of Regulation (EC) No 1907/2006, read-across from a structurally related substance is conducted.

In accordance with Article 13 (1) of Regulation (EC) No 1907/2006, "information on intrinsic properties of substances may be generated by means other than tests, provided that the conditions set out in Annex XI are met.” In particular for human toxicity, information shall be generated whenever possible by means other than vertebrate animal tests, which includes the use of information from structurally related substances (grouping or read-across).

Having regard to the general rules for grouping of substances and read-across approach laid down in Annex XI, Item 1.5, of Regulation (EC) No 1907/2006 whereby substances may be predicted as similar provided that their physicochemical and toxicological properties are likely to be similar or follow a regular pattern as a result of structural similarity.

Overview of Repeated dose toxicity via the oral route.

#	EC 905-964-4	CAS 102-76-1
Chemical name	"Reaction mixture of glycerol-1,3-di(acetate), glycerol acetate and triacetin"	Triacetin
Molecular weight	134.13 - 218.20 g/mol	218.20 g/mol
Repeated dose toxicity, oral	RA: CAS 102-76-1	NOAEL ≥ 1000 mg/kg bw/day (m,f)

 

The above mentioned substances are considered to be similar on the basis of the structural similarity resulting in similar properties and/or activities. The available endpoint information is used to predict the same endpoints for "Reaction mixture of glycerol-1,3-di(acetate), glycerol acetate and triacetin".

A detailed analogue approach justification is provided in the technical dossier (see IUCLID Section 13).

No studies are available investigating the repeated dose toxicity of "Reaction mixture of glycerol-1,3-di(acetate), glycerol acetate and triacetin".

In order to fulfil the standard information requirements set out in Annex IX, 8.6.2, in accordance with Regulation (EC) No 1907/2006 Annex XI, 1.5 read-across from the structurally related analogue substance Triacetin (CAS 102-76-1) is conducted.

A combined repeated dose toxicity study with the reproduction / developmental toxicity screening test was performed with Triacetin (CAS 102-76-1) according to OECD Guideline 422 and under GLP conditions (JECDB, 1998). Groups of twelve CD rats received oral gavage doses of 40, 200 and 1000 mg/kg bw/day of Triacetin (vehicle: 3% gum arabicum in purified water) for 44 days from 2 weeks prior to mating (males) and for 41-48 days from 14 days before mating to Day 3 postpartum (females). Concurrent control animals received the vehicle only. Triacetin had no effects on clinical signs, body weight, food consumption, and organ weight or necropsy findings. No abnormalities in haematological or blood chemical parameters in males were found. No histopathological changes were observed in either sex. Only one male animal of the high dose group died 32 days after start of the study. The death was not considered to be treatment related. The NOAEL for repeated dose oral toxicity was thus considered to be ≥ 1000 mg/kg bw/day for both sexes.

A 90-day oral feeding study in Sprague-Dawley rats was performed with a mixture of glycerol and Triacetin (CAS 102-76-1) to investigate nutritional needs during long-duration space missions (Shapira et al., 1969). Triacetin was mixed at concentrations of 16 and 30% to the diet. Groups of 8 male weanling Sprague-Dawley rats were fed the diet ad libitum for a period of up to 90 days. Assuming an average daily food consumption of 50 g/kg bw this approximates to 8000 and 15000 mg/kg bw/day. Control animals received the plain diet only. The authors concluded that growing rats could tolerate up to 20% Triacetin in the diet whereas higher amounts caused a decrease in weight gain. This corresponds to a NOAEL of 10000 mg/kg bw/day for Triacetin in male rats based on clinical signs and body weight gain.

In another study, glycerol, propylene glycol and Triacetin (CAS 102-76-1) were used as starch substitutes in young and adult rats (Shapira et al., 1975). The experiments were performed in order to evaluate nutritional needs associated with long term space missions. Triacetin was fed at concentrations of 20% and 30% of the starch-deficient diet (approximately 9000 and 15000 mg/kg bw/day) to groups of eight young and adult male Sprague-Dawley rats for a period of 13 weeks. Control animals received a diet containing starch only. Compared to the control animals, animals receiving Triacetin containing diets showed a reduced gain in body weight and increased mean liver weights. No mortalities occurred during the study period. The results are difficult to interpret, as the treated animals received a starch-deficient diet, which is probably a not suitable diet for rats. Therefore, a NOAEL could not be identified since the observed effects could be due to the diet-deficiency rather than the test substance treatment.

A 60 day oral feeding study with Triacetin (CAS 102-76-1) was performed in male and female rats to analyse the ability of fatty components to supply energy and support growth (Cox, 1933). Four rats, two males and two females were fed a diet containing 55% Triacetin (ad libitum) for 60 days. Triacetin at concentrations of 55% in the diet (supply of 77% of the caloric intake) supported normal growth and body weight gain in rats. Since no concurrent control animals fed with a normal rat diet were reported, no further evaluation of the results is possible.

Conclusion for Repeated Dose Toxicity – Oral

A Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test conducted with Triacetin (CAS 102-76-1) did not show effects in rats. Therefore, the NOAEL for repeated oral toxicity was found to be greater than 1000 mg/kg bw/day. In addition, Triacetin studies were available performed with rats to determine the nutritional needs during long-duration space missions. A 90-day feeding study with rats resulted in a well-tolerated amount of 20% Triacetin in diet (~10000 mg/kg bw/day). Overall, a NOAEL exceeding 1000 mg/kg bw for "Reaction mixture of glycerol-1,3-di(acetate), glycerol acetate and triacetin" was deduced.

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

Justification for classification or non-classification

Based on read-across from the source substance Triacetin following an analogue approach, the available data on repeated dose toxicity of "Reaction mixture of glycerol-1,3-di(acetate), glycerol acetate and triacetin" do not meet the criteria for classification

according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and the data are therefore conclusive but not sufficient for classification.