Androst-4-ene-3,17-dione

Endpoint summary

Description of key information

in vitro skin irritation: not irritating (Wingenroth, 2021)

in vivo skin irritation: not irritating (Kurth, 1996)

 

in vivo eye irritation: not irritating (Treher, 1995)

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

19 Oct 2020 - 22 Oct 2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

Version / remarks:

2020

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)

Version / remarks:

2019

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: no data available

Justification for test system used:

accordign to guideline

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: reconstructed human epidermis model epiCS® (Cat.-No.CS-1001, CellSystems, Troisdorf, Germany)
- Tissue batch number(s): 100·AJ0414-1
- Date of initiation of testing: 19-10-2020

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: RT (room temperature)
- Temperature of post-treatment incubation (if applicable): Incubator temperature: 37 ± 2° C (CO2 gas concentration: 5 %; Humidity: 95%)

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1mg/ml
- Incubation time: 3 hours
- Spectrophotometer: EL808, Bio-Tek
- Wavelength: 570 nm

NUMBER OF REPLICATE TISSUES: 3

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
- Killed control and Color control were not required.

NUMBER OF TESTING RUNS / EXPERIMENTS TO DERIVE FINAL PREDICTION: the optical density of the isopropanol-extracts of 3 insert was determined by duplicate per insert = 6 OD values.

PREDICTION MODEL / DECISION CRITERIA:

- The mean optical density (OD) values obtained with the test item were used to calculate the percentage of viability relative to the negative control, which is set at 100 %.
- according to UN GHS (Category 2 or Category 1) if the mean percent tissue viability after exposure and post treatment incubation is less than or equal (≤ ) to 50 %.


Acceptance criteria:
The following acceptance criteria determined the validity of an assay:
- mean OD negative control >/=0.8 and - mean relative viability of the positive control is - If the mean viability of the 3 replicates > 20% the coefficient of variation (CV) should not exceed 0.3.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 30 mg (plus 30 µl 0.9% NaCl to moisten and ensure good contact with the epidermis surface)

NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 30 µl
- Concentration (if solution): 0.9% NaCl in water

POSITIVE CONTROL
- Amount(s) applied (volume or weight): 30 µl
- Concentration (if solution): 5% SDS in physiological saline

Duration of treatment / exposure:

20 min

Duration of post-treatment incubation (if applicable):

42 h

Number of replicates:

3

Irritation / corrosion parameter:

% tissue viability

Value:

114.89

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

- OTHER EFFECTS:
- Direct-MTT reduction: no
- Colour interference with MTT: no

- DEMONSTRATION OF TECHNICAL PROFICIENCY:
Reliability of the test was previously confirmed by interlaboratory validation

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: yes
- Acceptance criteria met for positive control: yes
- Acceptance criteria met for variability between replicate measurements: yes

Conclusions:

A study was performed for the assessment of the skin irritancy of the test item Androstendione with reconstructed human epidermis (RhE) in accordance with OECD TG 439 and EU Test Method B.46.
The mean value of cell viability was recorded to be 114.89%. The test item was thus shown to be not irritating to reconstructed human skin in vitro.

Executive summary:

A study was performed for the assessment of the skin irritancy of the test item Androstendione with reconstructed human epidermis (RhE). The experiment was carried out in vitro using the commercially available test method epiCS®. The study was conducted in accordance with OECD TG 439 and EU Test Method B.46.

The test item was applied undiluted topically to the RhE tissue construct in triplicates and incubated for 20 minutes, followed by a 42 hours post-treatment incubation period.

Cell viability was measured in a photometer by the amount of MTT (methylthiazole tetrazolium) reduction. The optical density value obtained for the test item was used to calculate the percentage of viability relative to the negative control, which is set at 100%.

The results of the concurrent negative control (NC, 0.9 % NaCl) and positive control (PC, 5 % SDS) demonstrated the viability (NC) and sensitivity (PC) of the test model.

The following value of cell viability was recorded for the test item: 114.89%.

In conclusion the results of the assay used show no skin irritant properties of the test item  Androstendion.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

March 1995

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Version / remarks:

adopted 24 Feb. 87

Deviations:

no

GLP compliance:

yes

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Sex: females
- Source: Schriever
- Age at study initiation: not reported
- Weight at study initiation: 3.4-4.4 kg
- Housing: singly in conventional housing conditions (metal cage)
- Diet and water: ad libitum
- Acclimation period: > 14 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22
- Humidity (%): 48-56
- Air changes (per hr): not reported
- Photoperiod (hrs dark / hrs light): 12/12

Vehicle:

unchanged (no vehicle)

Controls:

other: the left eye remained untreated and served as control

Amount / concentration applied:

0.1 mL, corresponding to 54.0-64.3 mg substance

Duration of treatment / exposure:

0.1 mL substance was applied into the conjunctival sac of the right eye; no rinsing of the eyes reported.

Observation period (in vivo):

Examinations were terminated after 4 days, since no changes at all were observed during this period.

Number of animals or in vitro replicates:

4 (females)

Details on study design:

One animal was treated in advance of the others, to ensure that if a severe response was produced, no further animals would be exposed. 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.

SCORING SYSTEM:
According to "Illustrated Guide for Grading Eye Irritation by Hazardous Substances", US Department of Health, Education and Welfare, Food and Drug Administration, Washington DC 20204, USA.

Irritation parameter:

cornea opacity score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

cornea opacity score

Basis:

animal #4

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

iris score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

iris score

Basis:

animal #4

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

3

Irritation parameter:

conjunctivae score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

3

Irritation parameter:

conjunctivae score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

3

Irritation parameter:

conjunctivae score

Basis:

animal #4

Time point:

24/48/72 h

Score:

0

Max. score:

3

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

chemosis score

Basis:

animal #4

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritant / corrosive response data:

The single application of the substance into the conjunctival sac of the rabbit eye led to slight to moderate transient irritation (vessel injection and secreation in 2/4 animals) solely on the day of application. From day 2 onwards all animals were without findings.

Other effects:

not reported

Conclusions:

The test item was tolerated without any local findings in 2 of 4 animals. Only one animal each showed slight injection of blood vessels in the conjunctiva palpebrae and moderate secretion, respectively, on application day. From day 2 onwards, both animals were without findings as well. In this study, Androstendion was not irritating to the eye.

Executive summary:

In a primary eye irritation study according to OECD TG 405 0.1 ml Androstendione (corresponding to 54.0 to 64.3 mg) was instilled into the conjunctival sac of the right eye of New Zealand White rabbits (4 females). The untreated left eye served as control. Animals were observed for 4 d. 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.

The test item was tolerated without any local findings in 2 of 4 animals. Only one animal each showed slight injection of blood vessels in the conjunctiva palpebrae and moderate secretion, respectively, on application day. From day 2 onwards, both animals were without findings as well. The control eyes were without findings. In this study, Androstendion was not irritating to the eye.  

 

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation

A study was performed for the assessment of the skin irritancy of the test item Androstendion with reconstructed human epidermis (RhE). The experiment was carried out in vitro using the commercially available test method epiCS®. The study was conducted in accordance with OECD TG 439 and EU Test Method B.46.

The test item was applied undiluted topically to the RhE tissue construct in triplicates and incubated for 20 minutes, followed by a 42 hours post-treatment incubation period.

Cell viability was measured in a photometer by the amount of MTT (methylthiazole tetrazolium) reduction. The optical density value obtained for the test item was used to calculate the percentage of viability relative to the negative control, which is set at 100%.

The results of the concurrent negative control (NC, 0.9 % NaCl) and positive control (PC, 5 % SDS) demonstrated the viability (NC) and sensitivity (PC) of the test model.

The following value of cell viability was recorded for the test item: 114.89%.

In conclusion the results of the assay used show no skin irritant properties of the test item  Androstendion (Wingenroth, 2021).

 

In a combined study on acute toxicity and on local tolerance similar to OECD TG 402 Wistar rats (3/sex) were dermally exposed to Androstendion in physiological saline for 24 hours at a limit dose of 2000 mg/kg bw under semiocclusive 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

In a primary eye irritation study according to OECD TG 405 0.1 ml Androstendion (corresponding to 54.0 to 64.3 mg) was instilled into the conjunctival sac of the right eye of New Zealand White rabbits (4 females). The untreated left eye served as control. Animals were observed for 4 d. 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.

The test item was tolerated without any local findings in 2 of 4 animals. Only one animal each showed slight injection of blood vessels in the conjunctiva palpebrae and moderate secretion, respectively, on application day. From day 2 onwards, both animals were without findings as well. The control eyes were without findings. In this study, Androstendion was not irritating to the eye (Treher, 1995).  

 

No study investigating a respiratory irritant potential of the substance is available. But as no irritant potential was seen in the above specified in vivo irritation/corrosion studies on skin and eye (all scores zero at 24, 48, and 72 h), it is unlikely that the substance would show a respiratory irritant property. 

Justification for classification or non-classification

No classification required for skin, eye or respiratory irritation/corrosion according to Regulation (EC) 1272/2008.