1,4-Dioxane-2,5-dione, 3,6-dimethyl-, (3R,6R)-

Endpoint summary

Description of key information

A sample of D-lactide was examined for acute oral toxicity in an experiment conducted according to OECD guideline 423 with female rats. A dose level of 2000 mg/kg body weight was examined. No mortality or distinct clinical signs were observed after treatment of 6 females with the 2000 mg/kg bw dose level.
In an acute dermal toxicity study (limit test), a group of young adult Wistar rats (5 males and 5 females) was dermally exposed to D-lactide in polyethylene glycol 400 for 24 hours to approximately 10% of body surface area at a dose level of 2000 mg/kg bw. Animals were observed for 14 days. No mortality occurred. There were no treatment related clinical signs, necropsy findings or changes in body weight.
Lactic acid is used as a read-across partner for D-lactide and in an acute inhalation toxicity study in rats with lactic acid a LC50 of  > 7.94 mg/L air was determined. Based on the results, no classification for acute toxicity is warranted.

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2010-05-28 to 2010-09-27

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)

Deviations:

no

GLP compliance:

yes

Test type:

acute toxic class method

Limit test:

yes

Specific details on test material used for the study:

- Name of test material (as cited in study report): D-Lactide
- Batch: 0912000436
- Purity: 99%
- Expiration date of the lot/batch: 2010-12-04
- Stability under test conditions: stable
- Storage condition of test material: in refrigerator (2-8 °C) in the dark under nitrogen

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany.
- Age at study initiation: approximately 8 weeks old.
- Weight at study initiation:Body weight variation did not exceed ± 20 % of the sex mean (females: 159 g).
- Fasting period before study: Animal were deprived of food overnight prior to dosing and until 3-4 hours after administration of the test substances. Water was available.
- Housing: Group housing of 3 animals per cage inlabeled Macrolon cages (MIV type; height 18 cm) containing sterilized sawdust as bedding material (Litalabo, S.P.P.S., Argenteuil, France) and paper as cage-enrichment (Enviro-dri, Wm. Lillico & Son (Wonham Mill Ltd), Surrey, United Kingdom).
- Diet (e.g. ad libitum): Free access to pelleted rodent diet (SM R/M-Z from SSNIFF@Spezialdiäten GmbH, Soest, Germany).
- Water (e.g. ad libitum): Free access to tap water.
- Acclimation period: At least 5 days before start of treatment under laboratory conditions.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.0 ± 3.0 (actual range: 19.8 - 21.5)
- Humidity (%): Relative humidity of 40-70 (actual range: 38-75)
- Air changes (per h): Approximately 15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours artificial fluorescent light and 12 hours darkness per day

IN-LIFE DATES: From 04 June 2010 to: 23 June 2010

Route of administration:

oral: gavage

Vehicle:

polyethylene glycol

Details on oral exposure:

VEHICLE
- Dose level (volume): 2000 mg/kg bw (10 mL/kg bw)
- Justification for choice of vehicle: The vehicle was selected based on trial formulations performed at NOTOX and on test substance data supplied by the sponsor.

MAXIMUM DOSE VOLUME APPLIED: 2000 mg/kg bw (10 mL/kg bw)

DOSAGE PREPARATION (if unusual): The vehicle was dehydrated before the formulations were prepared. The formulations (w/w) were prepared within 4 hours prior to dosing. Homogeneity was accomplished to a visually acceptable level. Adjustment was made for specific gravity of the vehicle. In order to obtain homogeneity, the test substance formulations were heated in a water bath with a maximum temperature of 50 °C for a maximum of 31 minutes. The test substance formulations were allowed to cool down below 40 °C prior to dosing.

CLASS METHOD (if applicable)
- Rationale for the selection of the starting dose: Not given in the report.

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

3 females per dose (same dose tested twice, so 6 females in total)

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Mortality/viability: Twice daily. Body weights: Days 1 (pre-administration), 8 and 15.
- Necropsy of survivors performed: yes
- Other examinations performed: Clinical signs: At periodic intervals on the day of dosing (day 1) and once daily thereafter, until day 15. The time of onset, degree and duration were recorded and the symptomgraded according to fixed scales.

Statistics:

N.A.

Key result

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortality occurred.

Clinical signs:

other: Hunched posture was noted for all animals on day 1. In addition, lethargy and piloerection were noted among the majority of animals on day 1.

Body weight:

other body weight observations

Remarks:

No adverse effects observed

Gross pathology:

No abnormalities were found at macroscopic post mortem examination of the animals.

Other findings:

N.A.

Interpretation of results:

other: CLP criteria not met

Conclusions:

Since no mortality occurred during the 14-day observation period, the LD50 of D-lactide exceeds 2000 mg/kg bw in female rats. Therefore, D-lactide is considered as not harmful upon ingestion.

Executive summary:

In an acute oral toxicity study conducted according to OECD guideline 423, group of male and female Wistar rats (n= 3/sex) were given a single oral dose of D-lactide (purity >99%) at dose level of 2000 mg/kg body weight.

No mortality or distinct clinical signs, except hunched posture, piloerection and lethargy on day 1, were observed after treatment of 6 females with the 2000 mg/kg bw dose level. Macroscopic examination of the animals at the end of the observation period did not reveal any treatment-related gross changes. Since no mortality occurred during the 14-day observation period, the oral LD50 of D-lactide is considered to exceed 2000 mg/kg bw in female rats. D-lactide is considered as not harmful upon ingestion.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

> 2 000 mg/kg bw

Quality of whole database:

GLP guideline study

Acute toxicity: via inhalation route

Link to relevant study records

Reference

Endpoint:

acute toxicity: inhalation

Type of information:

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

Adequacy of study:

key study

Justification for type of information:

For details and justification of read-across please refer to the read-across report attached to IUCLID section 13.

Reason / purpose for cross-reference:

read-across source

Duration of exposure:

h

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 7.94 mg/L air

Based on:

test mat.

Exp. duration:

4 h

Mortality:

1 female died.

Clinical signs:

other: Please refer to box "Any other information on results incl. tables".

Body weight:

At the beginning of the study, mean body weicihts for individual groups were within 20% of the overall mean for each sex. All groups of male rats gained weight within the first week after exposure in comparison to pre-exposure weights (3% for sham-exposed, 2< for L(+)-lactic acid,respectively). Female rats in the sham group gained weight during the first week after exposure (less than 1%). Female rats in the treated group lost weight during the first week after exposure (7%). After 14 days, all surviving animals had gained weight in comparison to pre-exposure weights (14% for males, 7% for females). No significant differences were observed in body weight between treated and control groups.

Gross pathology:

All surviving animals were necropsied at the termination of the study. The animal that died during the study was necropsied immediately. No gross lesions were observed at necropsy.

Clinical signs:

Rapid breathing and eye tearing were observed during exposure. At one and three hours after exposure, all animals (including the sham controls) had a hunched posture, ruffled and ungroomed fur, brown stained fur and red-stained fur surrounding the eyes (tearing). By 24 hours, female treated rats had ruffled and stained coats. All other animals appeared normal at 24 hours and for the remainder of the 14 day observation period. Several treated female rats continued to have ruffled fur up to 4 days after exposure.

Mass Median Diameter:

The Mass Median Diameters ranged from 2.03 to 2.14 microns and averaged 2.09.

Interpretation of results:

other: CLP criteria not met

Conclusions:

Based on these results, the LC50 of L(+)-lactic acid is greater than 7.94 mg/L.

Executive summary:

In an acute inhalation toxicity study according to OECD 403, groups of young adult F344 rats (5/sex/dose) were exposed by inhalation route to L(+)-lactic acid in a concentration of approximately 7.94 mg/L for 4 hours.

Rapid breathing and eye tearing were observed during exposure. At one and three hours after exposure, all animnals (including the sham controls) had a hunched posture, ruffled and ungroomed fur, brown stained fur and red-stained fur surrounding the eyes (tearing). After 24 hours, female treated rats had ruffled and stained coats. All other animals appeared normal at 24 hours and for the remainder of the 14 day observation period. Several treated female rats continued to have ruffled fur up to 4 days after exposure. One female rat from the treated group died on day 9. All other animals survived until the end of the study. At gross pathology no adverse lesions were observed.

Based on these results, the LC50 of L(+)-lactic acid is greater than 7.94 mg/L.

This acute inhalation study is classified as acceptable. It does satisfy the guideline requirement for an acute inhaltion study (OECD TG 403) in rats.

This information is used in a read-across approach in the assessment of the target substance. For details and justification of read-across please refer to the read-across report attached to IUCLID section 13.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LC50

Value:

> 7 940 mg/m³ air

Physical form:

inhalation: aerosol

Quality of whole database:

Guideline study

Acute toxicity: via dermal route

Link to relevant study records

Reference

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2010-05-28 to 2010-09-27

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Version / remarks:

1987

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.3 (Acute Toxicity (Dermal))

Deviations:

no

GLP compliance:

yes

Test type:

standard acute method

Limit test:

yes

Specific details on test material used for the study:

- Name of test material (as cited in study report): D-Lactide
- Batch no.: 0912000436
- Appearance: white flakes
- Purity: 99%
- Expiration date of the lot/batch: 2010-12-04
- Stability under storage conditions: stable
- Storage condition of test material: in refrigerator (2-8 °C) in the dark under nitrogen

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany.
- Age at study initiation: approximately 11 weeks old.
- Weight at study initiation:Body weight variation did not exceed ± 20 % of the sex mean (males: 312 g, females: 217 g).
- Fasting period before study:not applicable.
- Housing: Individually housed in labeled Macrolon cages (MIII type; height 18 cm) containing sterilised sawdust as bedding material (Litalabo, S.P.P.S., Argenteuil, France) and paper as cage-enrichment (Enviro-dri, Wm. Lillico & Son (Wonham Mill Ltd), Surrey, United Kingdom).
- Diet (e.g. ad libitum): Free access to pelleted rodent diet (SM R/M-Z from SSNIFF@Spezialdiäten GmbH, Soest, Germany).
- Water (e.g. ad libitum): Free access to tap water.
- Acclimation period: At least 5 days before start of treatment under laboratory conditions. During the acclimatization period the animals were group housed in Macrolon cages (MIV type).

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.0 ± 3.0 (actual range: 19.8-21.5)
- Humidity (%): relative humidity of 40-70 (actual range: 38-75)
- Air changes (per h): approximately 15
- Photoperiod (hrs dark/hrs light): 12 / 12

IN-LIFE DATES: From 09 June 2010 to 23 June 2010

Type of coverage:

occlusive

Vehicle:

polyethylene glycol

Details on dermal exposure:

TEST SITE
- Area of exposure: Approximately 10% of the total body surface, i.e. approximately 25 cm² for males and 18 cm² for females.
- % coverage: Approximately 10% of the total body surface.
- Type of wrap if used:The test substance formulation was held in contact with the skin with a dressing, consisting of a surgical gauze patch (Surgy 1D, Laboratoires Stella s.a., Liege, Belgium), successively covered with aluminium foil and Coban elastic bandage (3M, St. Paul, Minnesota, U.S.A. (Coban & Micropore)).

REMOVAL OF TEST SUBSTANCE
- Washing (if done): The skin was cleaned of residual test substance using tap water.
- Time after start of exposure: 24 hours.

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 2000 mg/kg bw (10 mL/kg bw)

VEHICLE
Polyethylene glycol 400 (Merck, Darmstadt, Germany) (specific gravity 1.125). The vehicle was dehydrated before the formulation was prepared. The formulation (w/w) was prepared within 4 hours prior to dosing. Homogeneity was accomplished to a visually acceptable level. Adjustment was made for specific gravity of the vehicle. In order to obtain homogeneity, the test substance formulation was heated in a water bath with a temperature of 49.3 °C for 31 minutes. The test substance formulation was allowed to cool down below 40 °C prior to dosing.

Duration of exposure:

24 hours

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

5

Control animals:

not required

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Mortality/viability: Twice daily. Body weights: Days 1 (pre-administration), 8 and 15.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs: At periodic intervals on the day of dosing (day 1) and once daily thereafter, until day 15. The time of onset, degree and duration were recorded and the symptomgraded according to fixed scales.

Statistics:

N.A.

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortality occurred.

Clinical signs:

other: Chromodacryorrhoea (left eye) was noted for one male on day 2. One male showed scabs on day 6 and scales on day 7 in the treated skin area.

Body weight:

other body weight observations

Remarks:

No adverse effects were observed

Gross pathology:

No abnormalities were found at macroscopic post mortem examination of the animals.

Interpretation of results:

other: CLP criteria not met

Conclusions:

The dermal LD50 value of D-lactide in Wistar rats was established to exceed 2000 mg/kg body weight.

Executive summary:

In an acute dermal toxicity study conducted according to OECD guideline 402, a group of young adult Wistar rats (5 males and 5 females) was dermally exposed to D-lactide (purity 99%) in polyethylene glycol 400 for 24 hours to approximately 10% of body surface area at 2000 mg/kg bw. Animals were observed for 14 days. No mortality occurred. There were no treatment related necropsy findings or changes in body weight. Clinical signs as e.g. chromodacryorrhoea (left eye) was noted in one male on day 2 and one male shoed scabs on day 6 and scales on day 7 in treated skin area.

The dermal LD50 value of D-lactide in Wistar rats was established to exceed 2000 mg/kg body weight.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

> 2 000 mg/kg bw

Quality of whole database:

GLP guideline study

Additional information

After oral application of rats with D-lactide (study conducted in accordance with OECD 423) an acute oral LD50 of D-lactide of > 2000 mg/kg bw was determined. After dermal application (study conducted in accordance with OECD guideline 402), the LD50 of D-lactide was determined with > 2000 mg/kg bw.

Lactic acid is an acceptable read-across partner as lactide is rapidly converted by hydrolysis into lactic acid under aqueous conditions.

In an acute inhalation toxicity study in rats with lactic acid conducted in accordance with OECD guideline 403, a LC50 of > 7.94 mg/L air was determined.

Justification for classification or non-classification

Based on the available data and according to CLP Regulation 1272/2008, D-lactide does not warrant classification for acute toxicity. LD50 values for all routes are above the limit values of the relevant OECD guidelines.