Reaction mass of (2R*,4R*,4aR*,9bS*)-2,4-dimethyl-4,4a,5,9b-tetrahydroindeno[1,2-d][1,3]dioxine and (2R*,4R*,4aS*,9bR*)-2,4-dimethyl-4,4a,5,9b-tetrahydroindeno[1,2-d][1,3]dioxine

Endpoint summary

Description of key information

Oral (OECD 423), rat: LD50: 300 - 500 mg/kg bw
Dermal (OECD 402), rat: LD50 > 2000 mg/kg bw (limit test)

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:

07 Mar - 29 Mar 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP-Guideline study

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

adopted 22 March 1996

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

The Department of Health of the Government of the United Kingdom

Test type:

standard acute method

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 8 weeks
- Weight at study initiation: males: 227 - 249 g; females: 201 - 230 g
- Fasting period before study: overnight
- Housing: animals were housed in groups of three by sex in solid-floor polypropylene cages with woodflakes
- Diet: Rat and Mouse Expanded Diet No. 1 (Special Diet Services Limited, Witham, UK), ad libitum
- Water: mains drinking water, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 - 25
- Humidity (%): 30 - 70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12 / 12

Route of administration:

oral: gavage

Vehicle:

arachis oil

Remarks:

Only for the 200 mg/kg bw dose level, the test material was prepared as solution in arachis oil. For the 2000 mg/kg dose level, the test material was used ad supplied.

Details on oral exposure:

VEHICLE
- Justification for choice of vehicle: test material did not dissolve or suspend in distilled water

MAXIMUM DOSE VOLUME APPLIED: 1.83 mL/kg bw (2000 mg/kg bw); 10 mL/kg bw (200 mg/kg bw)

Doses:

2000 mg/kg bw (step 1)
200 mg/kg bw (step 2)

No. of animals per sex per dose:

3 females at 2000 mg/kg bw
3 males and females at 200 mg/kg bw

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Animals were observed 0.5, 1, 2 and 4 hours after dosing and subsequently once daily for 14 days. Individual bodyweights were recorded prior to dosing and 7 and 14 days after treatment or at death.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight, gross pathology

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 300 - 500 mg/kg bw

Based on:

test mat.

Remarks on result:

other: according to Appendix 3c in OECD Guideline 423 (adopted Mar 1996)

Mortality:

Two animals treated with 2000 mg/kg bw were found dead one or three days after dosing. One animal treated with 2000 mg/kg bw was killed in extremis one day after dosing. No mortalities were observed at a dose level of 200 mg/kg bw until the end of the study.

Clinical signs:

other: Signs of systemic toxicity noted at a dose level of 2000 mg/kg bw were hunched posture, lethargy, decreased respiratory rate, gasping, laboured and noisy respiration, ataxia, ptosis, dehydration, increased lachrymation, hypothermia, body tremors, prostrat

Gross pathology:

Abnormalities noted at necropsy of animals that died or were killed in extremis during the study were haemorrhagic lungs, dark liver or patchy pallor of the liver, dark kidneys, haemorrhage and sloughing of the gastric mucosa, sloughing of the non-glandular region of the stomach and haemorrhagic small intestine. No abnormalities were noted at necropsy of animals that were killed at the end of the study.

Table 1. Results of the acute toxicity study.

Dose level (mg/kg bw)	Mortalities
	Male	Female
2000	-	3/3
200	0/3	0/3

Interpretation of results:

harmful

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

In this acute oral toxicity study in rats a LD50 value of > 300 - 500 mg/kg bw was found.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LD50

Value:

300 mg/kg bw

Quality of whole database:

The available information comprises an adequate and reliable study (Klimisch score 1), and is thus sufficient to fulfil the standard information requirements set out in Annex VII, 8.5, of Regulation (EC) No 1907/2006.

Acute toxicity: via inhalation route

Endpoint conclusion

Endpoint conclusion:

no study available

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:

01 - 15 Mar 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP-Guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Version / remarks:

adopted 24 February 1987

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

The Department of Health of the Government of the United Kingdom

Test type:

standard acute method

Limit test:

yes

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 8 weeks
- Weight at study initiation: males: 231 - 245 g; females: 201 - 228 g
- Housing: individually during the 24-hour exposure period and in groups of 5, by sex, for the remainder of the study
- Diet: Rat and Mouse Expanded Diet No. 1 (Special Diets Services Limited, Witham, UK), ad libitum
- Water: mains drinking water, ad libitum
- Acclimation period: minimum 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 - 25
- Humidity (%): 30 - 70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12 / 12

Type of coverage:

semiocclusive

Vehicle:

unchanged (no vehicle)

Details on dermal exposure:

TEST SITE
- Area of exposure: back and flanks
- % coverage: 10% of the total body surface area
- Type of wrap if used: surgical gauze with self-adhesive bandage

REMOVAL OF TEST SUBSTANCE
- Washing (if done): yes, with cotton wool moistened with a suitable solvent
- Time after start of exposure: 24 hours

TEST MATERIAL
- Amount applied: 1.83 mL/kg bw

Duration of exposure:

24 hours

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

5

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Animals were subjected for deaths or overt signs of toxicity 0.5, 1, 2 and 4 h after dosing and subsequently once daily for 14 days. After removal of the dressings and subsequently once daily for 14 days, the test sites were examined for evidence of primary irritation anc scored according to Draize. Individual body weights were recorded prior to application of the test substance and 7 and 14 days after treatment.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight, gross pathology

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No deaths were noted until the end of the observation period.

Clinical signs:

other: No signs of systemic toxicity were noted until the end of the 14-day observation period. No skin irritation was observed at all reading time points the values for erythema and edema was 0.

Gross pathology:

No abnormalities were noted at necropsy.

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

In this acute dermal toxicity study a LD50 value > 2000 mg/kg bw in male and female rats was found.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Quality of whole database:

The available information comprises an adequate and reliable study (Klimisch score 1), and is thus sufficient to fulfil the standard information requirements set out in Annex VIII, 8.5, of Regulation (EC) No 1907/2006.

Additional information

Oral

The acute oral toxicity of the test substance was assessed in a study according to OECD Guideline 423 and in compliance with GLP (2001). In a first step, a total dose of 2000 mg/kg bw of the undiluted test substance was administered to 3 female rats. Animals were observed 0.5, 1, 2 and 4 hours after dosing and subsequently once daily for 14 days. Individual body weights were recorded prior to dosing and 7 and 14 days after treatment. Macroscopic examination was performed in the end of the observation period at terminal sacrifice. Two animals treated with 2000 mg/kg bw were found dead one or three days after dosing and one animal was killed in extremis one day after dosing. In a second step, 3 male and 3 female rats were treated with 200 mg/kg bw of the test substance diluted in arachis oil. No mortalities were observed at this a dose level until the end of the study. Signs of systemic toxicity noted at 2000 mg/kg bw were hunched posture, lethargy, decreased respiratory rate, gasping, laboured and noisy respiration, ataxia, ptosis, dehydration, increased lachrymation, hypothermia, body tremors, prostration, chromodacryorrhea, loss of righting reflex, pilo-erection and splayed gait. Two animals treated with 2000 mg/kg bw were found comatose 4 hours or 1 day after dosing. Signs of systemic toxicity noted at a dose level of 200 mg/kg bw were hunched posture, lethargy, ataxia, decreased respiratory rate, laboured respiration, increased salivation and splayed gait. All surviving animals showed expected body weight gain over the study period. Abnormalities noted at necropsy of animals that died or were killed in extremis during the study were haemorrhagic lungs, dark liver or patchy pallor of the liver, dark kidneys, haemorrhage and sloughing of the gastric mucosa, sloughing of the non-glandular region of the stomach and haemorrhagic small intestine. No abnormalities were noted at necropsy of animals that were killed at the end of the study. Based on the results of this study, the LD50 value was determined to be in the range of 300 - 500 mg/kg bw in rats.

 

Dermal

The acute dermal toxicity of the test substance was assessed in a limit test performed in 5 male and 5 female Wistar rats according to OECD Guideline 402 and in compliance with GLP (2001). A single dose of 2000 mg/kg bw of the test substance was applied to the shorn skin of rats under semi-occlusive conditions for 24 hours. Animals were subjected for deaths or overt signs of toxicity 0.5, 1, 2 and 4 hours after dosing and subsequently once daily for 14 days. After removal of the dressings and subsequently once daily for 14 days, the test sites were examined for evidence of primary irritation and scored according to Draize. Individual body weights were recorded prior to application of the test substance and 7 and 14 days after treatment. Macroscopic examination was performed in the end of the observation period at terminal sacrifice. There was no mortality and no effects on body weight gain during the 14-day observation period. No signs of systemic toxicity and no signs of dermal irritation were observed. No abnormalities were noted at necropsy. The LD50 value for dermal toxicity is considered to be > 2000 mg/kg bw.

In a second study, the dermal toxicity was assessed in mice. Two male mice per dose were treated with 0.464, 0.681, 1.00 and 1.47 mL/kg bw (equivalent to 505, 741, 1088 and 1599 mg/kg bw based on a relative density of 1.088) test substance diluted in 0.8% aq. hydroxylpropylmethylcellulose. No mortality was observed at 0.464 mL/kg bw. One animal died at 0.681 and 1.00 mL/kg bw. Two animals died at 1.47 mL/kg bw. Mortality was observed within 2-5 days after application. The LD50 value in mice was calculated to be 0.82 mL/kg bw (equivalent to 892 mg/kg bw). Because of the limited documentation, limited number of test animals and since no information on purity was given the study was considered not reliable.

Justification for selection of acute toxicity – oral endpoint
The reliable GLP compliant OECD Guideline study was chosen.

Justification for selection of acute toxicity – inhalation endpoint
No study required since exposure of humans via inhalation is unlikely taking into account the physico-chemical properties and the intended uses of the substance.

Justification for selection of acute toxicity – dermal endpoint
The reliable GLP compliant OECD Guideline study was chosen.

Justification for classification or non-classification

The available data on acute oral toxicity of the test substance meet the criteria for Acute Tox. Cat 4 (H302) according to Regulation (EC) 1272/2008.

The available data on acute dermal toxicity of the test substance do not meet the criteria for classification according to Regulation (EC) 1272/2008, and are therefore conclusive but not sufficient for classification.