Estr-4-ene-3,17-dione

Endpoint summary

Description of key information

Norandrostendion is not irritating to the skin of rats (Lihui, 2020) or to the eyes of rabbits (Lei, 2020 & Treher,1996).

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

26 December 2019 - 16 April 2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

The additional test in skin irritation was performed based on the Guidance on Environmental Management Registration of New Chemical Substances (Order No. 12 of the MEE of the P.R. China). The in vivo test was the accepted method of the toxicological endpoint skin irritation/corrosion at the time for a New Chemical Substance Registration in China.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Version / remarks:

2015

Qualifier:

according to guideline

Guideline:

other: State Ministry of Environmental Protection of the People’s Republic of China. Guidelines of Chemical Testing (HJ/T 153-2004) Annex A 404 Acute Dermal Irritation/Corrosion Test.

GLP compliance:

yes

Species:

rabbit

Strain:

other: Japanese long-eared white rabbit

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Qingdao Kangda Biological Technology Co., Ltd.
- Weight at study initiation: 2.30-2.40 kg
- Housing: individually
- Diet (e.g. ad libitum): rabbit maintenance feed (batch no. 19101861) provided by Liaoning Changsheng Biotechnology Co., Ltd., ad libitum
- Water (e.g. ad libitum): drinking water, ad libitum
- Acclimation period: 19 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.03-23.70 °C
- Humidity (%): 40.71-67.80%
- Photoperiod (hrs dark / hrs light): 12/12

Type of coverage:

occlusive

Preparation of test site:

clipped

Vehicle:

other: moistened with water

Controls:

other: untreeated left side of the spinal column on the back of teh animal served as control

Amount / concentration applied:

0.5 g

Duration of treatment / exposure:

4 h

Observation period:

72 h

Number of animals:

3

Details on study design:

TEST SITE
- Area of exposure: Two areas of skin on both sides of the midline below the shoulder blade were selected (approximately 2.5 cm x 2.5 cm)

REMOVAL OF TEST SUBSTANCE
- Washing (if done): ultra-pure water and absorbent cotton were used to wash the skin at the test area
- Time after start of exposure: 4 h

OBSERVATION TIME POINTS
Immediately after patch removal, 1, 24, 48, 72 h

SCORING SYSTEM:
- Method of calculation: Draize scale
Dermal Irritation Response Score
Skin response Score
Erythema and eschar formation
No erythema 0
Slight erythema (barely perceptible) 1
Clearly visible, easily defined erythema 2
Moderate to severe erythema 3
Severe erythema (purplish-red) to eschar formation 4
Oedema formation
No oedema 0
Slight oedema (barely perceptible) 1
Slight oedema (skin raised, clear outline) 2
Moderate oedema (skin raised approximately 1 mm) 3
Severe oedema (raised more than 1 mm, extending beyond the area of exposure) 4

Irritation parameter:

erythema score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

edema score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritant / corrosive response data:

no abnormalities or skin reactions were observed immediately after the dressing was removed, and until 72 hours after the dressing was removed

Conclusions:

Under the conditions of this test, based on observations seen and the assessment system adopted, mean dermal erythema/oedema scores at 24 hours,
48 hours, and 72 hours after the dressings were removed were 0/0, 0/0, and 0/0. Based on the GHS classification criteria for dermal irritation/corrosion, Norandrostendione did not meet the classification criteria.

Executive summary:

In a dermal irritation study equivalent to OECD Guideline 404 three male Japanese long-eared white rabbits were dermally exposed to Norandrostendione. 0.5 g of the test sample was administered to the clipped dorsal skin on the right side of each animal and the dorsal skin on the left side served as the control. Exposure time was four hours. The animals were observed clinically immediately after the dressing was removed and at one hour, 24 hours, 48 hours and 72 hours.
Skin response was observed immediately after the dressing was removed and at one hour, 24 hours, 48 hours and 72 hours, and was scored based on the skin irritation response scoring criteria.

In all three animals no abnormalities were observed immediately after the dressing was removed, and until 72 hours after the dressing was removed.

Under the conditions of this test, based on observations seen and the assessment system adopted, mean dermal erythema/oedema scores at 24 hours,
48 hours, and 72 hours after the dressings were removed were 0/0, 0/0, and 0/0. Based on the GHS classification criteria for dermal irritation/corrosion, Norandrostendione did not meet the classification criteria.

 

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Reference 1

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

Nov 1995

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Version / remarks:

adopted 26 Feb. 1987

Deviations:

yes

Remarks:

4 instead of 3 animals

GLP compliance:

not specified

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Schriever
- Weight at study initiation: males: 3.0 kg, females: 2.8 - 3.1 kg
- Housing: 1/cage
- Diet (e.g. ad libitum): pell. Altromin® K, ad libitum
- Water (e.g. ad libitum): demineralized water, ad libitum
- Acclimation period: > 14 days; during this period a diet containing 0.03% sulfaquinoxaline was offered for 7 consecutive days as coccidiosis prophylaxis; the animals received Jacutin® emulsion into the meatus acusticus exterilljg for prophylaxis
against mite infection of the ear and were vaccinated against rabbit haemorrhagic disease (Arvilap®)

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 - 22°C
- Humidity (%): 66 - 70%
- Photoperiod (hrs dark / hrs light): 12/12

Vehicle:

unchanged (no vehicle)

Controls:

other: the left eye, which remained untreated, served as control.

Amount / concentration applied:

0.1 ml

Duration of treatment / exposure:

test substance remained in the eye (eye was not rinsed)

Observation period (in vivo):

6 days

Number of animals or in vitro replicates:

2 males, 2 females

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): no

SCORING SYSTEM:
Draize scale

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

iris score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

24/48/72 h

Score:

0.4

Max. score:

3

Reversibility:

fully reversible

Irritation parameter:

chemosis score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Other effects:

Animal no. 605F was sacrificed on day 3 of the study for humane reasons since it suffered from a paresis of the hind legs due to dislocation of sacral vertebra. This animal was without findings from day 2 onwards.

Table 1: Results of the study

 

		Irritant effects (score)
Animal no. (sex)	Irritation parameter	24 h	48 h	72 h	Mean scores
1 (M)	Cornea	0	0	0	0.0
	Iris	0	0	0	0.0
	Conjunctiva (reddening)	0	0	0	0.0
	Conjunctiva (swelling)	0	0	0	0.0
2 (M)	Cornea	0	0	0	0.0
	Iris	0	0	0	0.0
	Conjunctiva (reddening)	2	0	0	0.7
	Conjunctiva (swelling)	0	0	0	0.0
3 (F)	Cornea	0	0	0	0.0
	Iris	0	0	0	0.0
	Conjunctiva (reddening)	0	0	0	0.0
	Conjunctiva (swelling)	0	0	0	0.0
4 (F)	Cornea	0	0	0	0.0
	Iris	0	0	0	0.0
	Conjunctiva (reddening)	2	0	0	0.7
	Conjunctiva (swelling)	0	0	0	0

 

 

The control eyes were without findings.

 

Findings after treatment with the test substance were slight to severe secretion, slight to moderate reddening and swelling of the outer eye lids and eye lid closure on the application day. Furthermore, slight to mainly moderate reddening and swelling of the conjunctivae and slight to moderate injection of blood vessels were observed. Additionally, in one animal moderate reddening of the iris was noted 1 and 2 hours after application. On day 2, 2 animals still showed slight to moderate reddening and vascular injection of the conjunctivae. From day 2 or 3 onwards, all animals were without findings.

Conclusions:

In a local tolerance test on the rabbit eye according to OECD TG 405 a single application of the test substance into the conjunctival sac provoked transient mainly moderate signs of irritation. The findings were fully reversible by day 3 after administration. According to EU classification criteria need not be labelled as an eye irritant.

Executive summary:

In an eye irritation study according to OECD Guideline 405, four New Zealand White rabbits (2M, 2F) each received a volume of 0.1 ml Norandrostenedione (corresponding to 60.1 - 64.0 mg substance) into the conjunctival sac of the right eye. The left eye remained untreated and served as control.

During application and observation on the application day (first day of the test) the animals were kept in restraining cages for approximately 2.5 hours. No food or water was available during this time. Conjunctivae, eyelids, cornea and iris were evaluated before application, 0.5, 1 and 2 hours thereafter, and then once daily until termination of the study on day 4.

Findings after treatment with the test substance were slight to severe secretion, slight to moderate reddening and swelling of the outer eye lids and eye lid closure on the application day. Furthermore, slight to mainly moderate reddening and swelling of the conjunctivae and slight to moderate injection of blood vessels were observed. Additionally, in one animal moderate reddening of the iris was noted 1 and 2 hours after application. On day 2, 2 animals still showed slight to moderate reddening and vascular injection of the conjunctivae. From day 2 or 3 onwards, all animals were without findings.

According to EU classification criteria need not be labelled as an eye irritant.

Reference 2

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

29 December 2019 - 4 January 2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

The additional test in eye irritation was performed based on the Guidance on Environmental Management Registration of New Chemical Substances (Order No. 12 of the MEE of the P.R. China). The in vivo test was the accepted method of the toxicological endpoint eye irritation at the time for a New Chemical Substance Registration in China.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Qualifier:

according to guideline

Guideline:

other: State Ministry of Environmental Protection of the People’s Republic of China. Guidelines of Chemical Testing (HJ/T 153-2004)-405 Acute Eye Irritation/Corrosion Test.

GLP compliance:

yes (incl. QA statement)

Species:

rabbit

Strain:

other: Japanese long-eared white rabbit

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Qingdao Kangda Biological Technology Co., Ltd.
- Weight at study initiation: 2.30-2.35 kg
- Housing: individually
- Diet (e.g. ad libitum): rabbit maintenance feed provided by Liaoning Changsheng Biotechnology Co., Ltd. (batch no.: 19101861), ad libitum
- Water (e.g. ad libitum): every-day drinking water, ad libitum
- Acclimation period: 18 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.03-23.94 °C
- Humidity (%): 40.71-63.10%.
- Photoperiod (hrs dark / hrs light): 12/12

Vehicle:

unchanged (no vehicle)

Controls:

other: The untreated left eye served as the autologous control.

Amount / concentration applied:

0.1 g

Duration of treatment / exposure:

24 h

Observation period (in vivo):

72 h

Number of animals or in vitro replicates:

3 males

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): after 24 h the eyes were examited with fluorescein and rinsed with physiological saline

TOOL USED TO ASSESS SCORE: headlight magnifier / fluorescein

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

24/48/72 h

Score:

4

Max. score:

4

Irritation parameter:

iris score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

3

Irritation parameter:

chemosis score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritant / corrosive response data:

In the test period, animal nos. 1100, 1101, and 1102 did not exhibit retention of sodium fluorescein at 24 hours after test chemical administration.
In animal no. 1100, at one hour after test chemical administration right eye scores for cornea/iris/conjunctiva hyperaemia/chemosis were 0/0/0/0 and continued until 72 hours after test chemical administration. No other abnormal reactions were seen.
In animal no. 1101, at one hour after test chemical administration right eye scores for cornea/iris/conjunctiva hyperaemia/chemosis were 0/0/0/0, and continued until 72 hours after test chemical administration. No other abnormal reactions were seen.
In animal no. 1102, at one hour after test chemical administration right eye scores for cornea/iris/conjunctiva hyperaemia/chemosis were 0/0/0/0, and continued until 72 hours after test chemical administration. No other abnormal reactions were seen.
In animal nos. 1100, 1101, and 1102, mean scores for cornea/iris/conjunctiva
hyperaemia/chemosis at 24 hours, 48 hours, and 72 hours after test chemical administration were 0/0/0/0.

Other effects:

In the test period, all animals gained weight.

Conclusions:

In this study, mean scores for cornea/iris/conjunctiva hyperaemia/chemosis at 24 hours, 48 hours, and 72 hours after test chemical administration in the right eye
were 0/0/0/0. This test sample did not cause irritation to the eye in Japanese long-eared white rabbits. According to the GHS standard for categorising eye irritation and corrosion, the test sample was categorised as insufficient for categorisation.

Executive summary:

This test is carried out in accordance with the requirements of the State Ministry of Environmental Protection’s Guidelines of Chemical Testing (HJ/T 153-2004)-405, which is comparable to OECD Guideline 405. Japanese long-eared white rabbits were used as the test animals to perform the NAD acute eye irritation/corrosion test in rabbits.

Three male rabbits were used for the test and 0.1 g of the test sample was applied to the right eye of each animal with the untreated left eye serving as the autologous control. Eye irritation response was observed immediately after test chemical administration and at 1 hour, 7 hours, 24 hours, 30 hours, 48 hours, 54 hours, 72 hours, and 78 hours. The 24 hour, 48 hour, and 72 hour mean eye irritation scores were calculated.

In all animals (animal nos.: 1100, 1101, and 1102), the initial response after test chemical administration was that they closed their eyes. No abnormalities were seen one hour after test chemical administration and this situation continued until 78 hours after test chemical administration.
In the test period, animal nos. 1100, 1101, and 1102 did not exhibit retention of sodium fluorescein at 24 hours after test chemical administration.
In animal no. 1100, at one hour after test chemical administration right eye scores for cornea/iris/conjunctiva hyperaemia/chemosis were 0/0/0/0 and continued until 72 hours after test chemical administration. No other abnormal reactions were seen.
In animal no. 1101, at one hour after test chemical administration right eye scores for cornea/iris/conjunctiva hyperaemia/chemosis were 0/0/0/0, and continued until 72 hours after test chemical administration. No other abnormal reactions were seen.
In animal no. 1102, at one hour after test c hemical administration right eye scores for cornea/iris/conjunctiva hyperaemia/chemosis were 0/0/0/0, and continued until 72 hours after test chemical administration. No other abnormal reactions were seen.
In animal nos. 1100, 1101, and 1102, mean scores for cornea/iris/conjunctiva hyperaemia/chemosis at 24 hours, 48 hours, and 72 hours after test chemical administration were 0/0/0/0.

The mean scores for cornea/iris/conjunctiva hyperaemia/chemosis at 24 hours, 48 hours, and 72 hours after test chemical administration in
the right eye were 0/0/0/0. This test sample did not cause irritation to the eye of the Japanese long-eared white rabbit. According to the GHS standard for categorising eye irritation and corrosion, the test sample was categorised as insufficient for categorisation.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation

In a dermal irritation study equivalent to OECD Guideline 404 three male Japanese long-eared white rabbits were dermally exposed to Norandrostendione. 0.5 g of the test sample was administered to the clipped dorsal skin on the right side of each animal and the dorsal skin on the left side served as the control. Exposure time was four hours. The animals were observed clinically immediately after the dressing was removed and at one hour, 24 hours, 48 hours and 72 hours.
Skin response was observed immediately after the dressing was removed and at one hour, 24 hours, 48 hours and 72 hours, and was scored based on the skin irritation response scoring criteria.

In all three animals no abnormalities were observed immediately after the dressing was removed, and until 72 hours after the dressing was removed.

Under the conditions of this test, based on observations seen and the assessment system adopted, mean dermal erythema/oedema scores at 24 hours,
48 hours, and 72 hours after the dressings were removed were 0/0, 0/0, and 0/0. Based on the GHS classification criteria for dermal irritation/corrosion, Norandrostendione did not meet the classification criteria (Lihui, 2020).

 

In a combined study on acute toxicity and on local tolerance similar to OECD TG 402 and OECD TG 404 Wistar rats (3/sex) were dermally exposed to Norandrostendione in physiological saline for 24 hours at a limit dose of 2000 mg/kg bw under occlusive conditions.  Animals then were observed for 14 days.

The administration of the test substance was tolerated without mortalities, compound-related clinical findings, effects on body weight gain and gross pathological findings.

No local intolerance reactions at the application sites were observed (Kurth, 1996).

 

Eye irritation

This test is carried out in accordance with the requirements of the State Ministry of Environmental Protection’s Guidelines of Chemical Testing (HJ/T 153-2004)-405, which is comparable to OECD Guideline 405. Japanese long-eared white rabbits were used as the test animals to perform the NAD acute eye irritation/corrosion test in rabbits.

Three male rabbits were used for the test and 0.1 g of the test sample was applied to the right eye of each animal with the untreated left eye serving as the autologous control. Eye irritation response was observed immediately after test chemical administration and at 1 hour, 7 hours, 24 hours, 30 hours, 48 hours, 54 hours, 72 hours, and 78 hours. The 24 hour, 48 hour, and 72 hour mean eye irritation scores were calculated.

In all animals (animal nos.: 1100, 1101, and 1102), the initial response after test chemical administration was that they closed their eyes. No abnormalities were seen one hour after test chemical administration and this situation continued until 78 hours after test chemical administration.
In the test period, animal nos. 1100, 1101, and 1102 did not exhibit retention of sodium fluorescein at 24 hours after test chemical administration.
In animal no. 1100, at one hour after test chemical administration right eye scores for cornea/iris/conjunctiva hyperaemia/chemosis were 0/0/0/0 and continued until 72 hours after test chemical administration. No other abnormal reactions were seen.
In animal no. 1101, at one hour after test chemical administration right eye scores for cornea/iris/conjunctiva hyperaemia/chemosis were 0/0/0/0, and continued until 72 hours after test chemical administration. No other abnormal reactions were seen.
In animal no. 1102, at one hour after test c hemical administration right eye scores for cornea/iris/conjunctiva hyperaemia/chemosis were 0/0/0/0, and continued until 72 hours after test chemical administration. No other abnormal reactions were seen.
In animal nos. 1100, 1101, and 1102, mean scores for cornea/iris/conjunctiva hyperaemia/chemosis at 24 hours, 48 hours, and 72 hours after test chemical administration were 0/0/0/0.

The mean scores for cornea/iris/conjunctiva hyperaemia/chemosis at 24 hours, 48 hours, and 72 hours after test chemical administration in
the right eye were 0/0/0/0. This test sample did not cause irritation to the eye of the Japanese long-eared white rabbit. According to the GHS standard for categorising eye irritation and corrosion, the test sample was categorised as insufficient for categorisation (Lei, 2020).

 

In an eye irritation study according to OECD Guideline 405, four New Zealand White rabbits (2M, 2F) each received a volume of 0.1 ml Norandrostenedione (corresponding to 60.1 - 64.0 mg substance) into the conjunctival sac of the right eye. The left eye remained untreated and served as control.

During application and observation on the application day (first day of the test) the animals were kept in restraining cages for approximately 2.5 hours. No food or water was available during this time. Conjunctivae, eyelids, cornea and iris were evaluated before application, 0.5, 1 and 2 hours thereafter, and then once daily until termination of the study on day 4.

Findings after treatment with the test substance were slight to severe secretion, slight to moderate reddening and swelling of the outer eye lids and eye lid closure on the application day. Furthermore, slight to mainly moderate reddening and swelling of the conjunctivae and slight to moderate injection of blood vessels were observed. Additionally, in one animal moderate reddening of the iris was noted 1 and 2 hours after application. On day 2, 2 animals still showed slight to moderate reddening and vascular injection of the conjunctivae. From day 2 or 3 onwards, all animals were without findings.

According to EU classification criteria need not be labelled as an eye irritant (Treher, 1996).

Justification for classification or non-classification

Based on the study results a classification according to Regulation (EC) No. 1272/2008 (CLP) is not required.