Potassium dicyanoaurate

Endpoint summary

Description of key information

In vitro skin irritation and corrosion assessment in a reconstructed human epidermis model (EpiSkin) (OECD439).
In vivo skin irritation/corrosion test in the rabbit (OECD404).
In vitro eye irritation/corrosion test (BCOP) (OECD437).
In vivo eye irritation test in the rabbit (OECD405).

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Reference 1

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

30 October 2013 to 04 November 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliant, guideline test, available as an unpublished report.

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

The control groups were shared with study 41302738. The control raw data were filled with study 41302738. This deviation was considered not to affect the purpose or integrity of the study.

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

The control groups were shared with study 41302738. The control raw data were filled with study 41302738. This deviation was considered not to affect the purpose or integrity of the study.

GLP compliance:

yes (incl. QA statement)

Species:

other: EPISKIN (TM) reconstructed human epidermis model

Strain:

other: not applicable

Details on test animals or test system and environmental conditions:

Not applicable.

Type of coverage:

other: Topical

Preparation of test site:

other: not applicable

Vehicle:

unchanged (no vehicle)

Controls:

other: Not applicable.

Amount / concentration applied:

- Treatment group: Test carried out in triplicate. Approximately 10 mg of the test item was applied topically, ensuring an even covering, to the epidermis surface which had previously been moistened with 5 µL sterile, distilled water to improve contact between the solid test item and the epidermis.
- Negative control: Triplicate tissues treated with 10 µL Dulbecco's Phosphate Buffered Saline (DPBS) with Ca++ and Mg++
- Positive control: Triplicate tissues. 10 µL Sodium Dodecyl Sulphate (SDS) at 5% w/v aqueous solution spread over entire surface of the epidermis using a pipette tip with the process being repeated after 7 minutes.

Duration of treatment / exposure:

- Treatment period: 15 minutes
- Post-exposure incubation: At the end of the exposure period, tissues were rinsed with DPBS with Ca++ and Mg++. Rinsing was achieved by filling and emptying each tissue insert for approximately 40 seconds using a soft stream of DPBS to gently remove any residual test item. The rinsed tissues were transferred to a second column of three wells, each containing 2 mL of maintenance medium, and incubated for 42 hours, at 37°C and 5% CO2 in air.

Observation period:

Not applicable.

Number of animals:

Not applicable.

Details on study design:

Preincubation
2 mL of maintance medium, warmed to approximately 37°C, was pipetted into the first column of 3 wells of a pre-labeled 12-well plate. Each epiderimis unit was transferred into the maintance medium filled wells (3 units per plate). A different 12-well plate was used for the test item and each control item. The tissues were incubated at 37°C, 5% Co2 in air overnight.

Application of test Item and rinsing (Day 1)
2 mL of maintenance medium, warmed to approx. 37 °C, was pipetted into the second column of 3 wells of the 12-well plate. Triplicate tissues were treated with the test item for an exposure period of 15 minutes. The test item was applied topically to the coresponding tissues ensuring uniform covering. 5 µL sterile distilled water was topically applied to the epidermal surface in order to improve further contact between the test item and the epidermis. 10 mg of the test item was then applied to the epidermal surface. Triplicate tissues treated with 10 µL of DPBS served as the negative controls and triplicate tissues treated with 10 µL of SDS 5% w/v served as the positive controls. The plates were kept in the biological safety cabnet at room temperature for 15 minutes. At the end of the exposure period, each tissue was removed from the well using forceps and rinsed using a wash bottle containing DPBS with Ca ++ and Mg++. The rinsed tissues were transferred to the second column of 3 wells containing 2 mL of maintenance medium in each well. The rinsed tissue were incubated at 37°C., 5% CO2 in air for 42 hours.


The measurement of tissue viability (cytotoxicity) was measured by means of the colourimetric MTT reduction assay. Cell viability was measured by enzymatic reduction of the yellow MTT tetrazolium salt to a blue formazan salt (by the mitochondrial succinate dehdroganase in viable cells) in the test item treated tissue relative to the negative control using the following procedure. After incubation, each 12-well plate was placed onto a plate shaker for 15 minutes to homogenize the released mediators in the maintenance medium. 1.6 mL of the maintenance medium from beneath each tissue was transferred to pre-labeled micro tubes and stored in a freezer at -14 to -30°C for possible inflamatory mediator determination. 2 mL of a 0.3 mg/mL MTT solution, freshly prepared in assay medium, was pipetted into the third column of 3 wells of the 12-well plate(s). The tissues were transferred to the MTT filled wells, and were incubated for 3 hours at 37°C, 5% CO2 in air. Then the epidermis was carefully separated from the collagen matrix using forceps and both parts (epiderimis and collagen matrix) placed into labeled 1.5 mL micro tubes containing 500 µL of acidified isopropanol, ensuring that both the epidermis and collagen matrix were fully immersed. Each tube was plugged and mixed thoroughly on a vortex mixer. The tubes were refrigerated at 1 to 10° C until Day 6 of the experiment, allowing the extraction of formazan crystals out of the MTT-loaded tissues.

Irritation / corrosion parameter:

other: other: % relative viability

Value:

7.1

Remarks on result:

other:

Remarks:

Basis: mean. Time point: 15 minutes exposure followed by the 42-hour post-exposure incubation period. Reversibility: no data. Remarks: SD of relative mean vaibility (%) = 1.1. (migrated information)

Other effects / acceptance of results:

The relative mean viability of the test item treated tissues after 15-minute exposure period and 42 hours post-exposure incubation period was 7.1% with standard deviation of 1.1%.
The mean optical density measured at 562 nm was 0.067 with standard deviation of 0.011.
The test item is considered to be irritant using the EPISKIN (TM) human epidermis model.

- Positive control: The relative mean tissue viability was 10.4 % relative to the negative control and the standard deviation was 4.5 %.
- Negative control: The mean optical density was 0.950 and the standard deviation was 0.037.

The acceptance criteria were satisfied according to the protocol criteria for both the positive control (relative mean viability ≤ 40% and standard deviation ≤ 18%) and negative control (OD562≥ 0.6 and standard deviation of individual tissue viability ≤18%). The standard deviation of the triplicate treated tissues was 3.9 % and hence the test item acceptance criterion (≤ 18%) was also satisfied.

Table 1. Mean OD562 values and percentage viabilities for the negative control item, positive control item and test item.

Item	OD562of tisues	Mean OD562 of triplicate tissues	± SD of OD562	Relative individual tissue viability (%)	Relative mean viability (%)	± SD of relative mean viability (%)
Negative control item#	0.931	0.950	0.037	98.0	100*	3.9
	0.927			97.6
	0.993			104.5
Positive control item#	0.102	0.099	0.043	10.7	10.4	4.5
	0.054			5.7
	0.140			14.7
Test item	0.055	0.067	0.011	5.8	7.1	1.1
	0.071			7.5
	0.075			7.9

SD – standard deviation

*The mean viability of the negqative control tissues is set at 100%

OD₅₆₂– optical density

# - control group shared with Harlan study number 41302738

Interpretation of results:

Category 2 (irritant) based on GHS criteria

Conclusions:

Based on the relative mean viability of 7.1% the test item was classified as irritant. The following classification criteria apply: EU DSD (67/548/EEC) Irritant requires symbol 'Xi' risk phrase R38 'Irritant to skin'. EU CLP and UN GHS Hazard statement H315 'Causes Skin Irritation' Category 2.

Executive summary:

The purpose of the test was to evaluate the skin irritation potentail of the potassium dicyanoaurate using the EPISKIN reconstructed human epiderimis model according to the OECD 439 guideline. Triplicate tissues were treated with the test item for an exposure period of 15 minutes. At the end of the exposure period each tissue was rinsed before incubating for 42 hours. Results were the relative mean vaibility of the test item treated tissues was 7.1%. The test item was classified as irritant. The following classification criteria apply: EU DSD (67/548/EEC) Irritant requires symbol 'Xi' risk phrase R38 'Irritant to skin'. EU CLP and UN GHS Hazard statement H315 'Causes Skin Irritation' Category 2.

Reference 2

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

25th February 1992 to 17th March 1992

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP-compliant, guideline study, available as an unpublished report.

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.4 (Acute Toxicity: Dermal Irritation / Corrosion)

Deviations:

no

GLP compliance:

yes

Species:

rabbit

Strain:

other: White russian (Albino)

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: ASTA Medica AG. D-4802 Halle-Künsebeck
- Age at study initiation: Males: 5 months; Female: 7 months
- Weight at study initiation: Males: 2.10, 2.53 kg; Female: 2.42 kg
- Housing: Stainless steel cage with grating floor, type ASTA, size: 48.5x40x36.5
- Diet (e.g. ad libitum): ssniff K, Special diet for rabbits. Approx: 120 g/day per animal.
- Water (e.g. ad libitum): ad libitum
- Acclimation period: One day

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.0 - 22.5°C
- Humidity (%): 50 - 60%
- Photoperiod (hrs dark / hrs light): 6 a.m. - 6 p.m. CET artificial lighting, 6 p.m. - 6 a.m. CET darkness.

Type of coverage:

occlusive

Preparation of test site:

shaved

Vehicle:

water

Remarks:

test substance moisetened to ensure tight contact.

Controls:

other: The opposite area to the treatment on each animal was given a blank treatment, acting as a control.

Amount / concentration applied:

0.5 g
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.5 g

VEHICLE (demineralised water)
- Amount(s) applied (volume or weight with unit): 0.14 mL

Duration of treatment / exposure:

4 hours

Observation period:

14 days

Number of animals:

3 (2 male, 1 female)

Details on study design:

TEST SITE
- Area of exposure:6.25 cm2 (patch size)
- Type of wrap if used: artifical wool coated with natural rubber layer and synthetic film glue layer. Bandage wrapped several times around the trunk (with 8cm) provided complete occlusion.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): Yes
- Time after start of exposure: 4 hours

SCORING SYSTEM: DRAIZE Grade

Irritation parameter:

other: Erythema and Eschar

Basis:

mean

Time point:

other: First 72 hours after exposure

Score:

2.33

Max. score:

4

Reversibility:

other: Disappeared in 2 out of the 3 animals by 14 days

Irritation parameter:

edema score

Basis:

mean

Time point:

other: First 72 hours after exposure

Score:

2.92

Max. score:

4

Reversibility:

other: Disappeared in 2 out of the 3 animals by 14 days.

Irritation parameter:

overall irritation score

Basis:

mean

Time point:

other: First 72 hours after exposure

Score:

5.3

Max. score:

8

Reversibility:

other: Effects disappeared in 2 out of the 3 animals by 14 days.

Remarks on result:

other: Sum of Erythema & Eschar and Edema score

Irritant / corrosive response data:

Severe changes in all animals

Other effects:

No systemic effects observed.

Table 1. Observed skin reactions to treatment

Reaction of the Exposed Skin	Hours/Days after Exposure	Draize Grade
		Animal 1	Animal 2	Animal 3
Erythema and Eschar	1 h	2	3	2
	24 h	2	3	2
	48 h	2	3	2
	72 h	2	3	2
	4 d	2	3	2
	7 d	2	3	2
	14 d	0	1	0
Edema	1 h	4	4	4
	24 h	2	3	3
	48 h	2	3	3
	72 h	1	3	3
	4 d	1	3	3
	7 d	1	3	2
	14 d	0	1	0

Interpretation of results:

Category 2 (irritant) based on GHS criteria

Remarks:

Criteria used for interpretation of results: Gilman et. al (1983), Relationship between the Primary Dermal Irritation Index and Occular Irritation, J. Toxicol.-Cut. & Ocular Toxicol. 2, 107-117 (1983)

Conclusions:

Observation of the skin of rabbits after dermal application of potassium dicyanoaurate showed that the substance is severely irriating to the skin of the rabbit. No systemic effects were observed.

Executive summary:

Observation of the skin of rabbits after dermal application of potassium dicyanoaurate produced a primary irritation index of 5.3. Therefore, the substance is classified as severely irriating to the skin of the rabbit. No systemic effects were observed.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Eye irritation

Link to relevant study records

Reference

Reference 1

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

from 06 November 2013 to 26 November 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliant, guideline test, available as an unpublished report. Reliable without restriction.

Qualifier:

according to guideline

Guideline:

OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants)

Deviations:

yes

Remarks:

The control group and other common data incl. eye receipt and preparation, technical documentation, reagent record and preparation, initial opacity readings, group allocation and optical density print out were shared with study 413027735.

Principles of method if other than guideline:

The 'open-chamber' method was adopted. The window-locking ring and glass were removed from the chamber prior to treatment. The 'open-chamber' method was also followed for the control groups although solutions were applied. Following application the ring and glass were replaced during the exposure period until rinsing.

GLP compliance:

yes (incl. QA statement)

Species:

other: Not applicable

Strain:

not specified

Details on test animals or tissues and environmental conditions:

Eyes from adult cattle were obtained from a local abattoir as a by-product from freshly slaughtered animals.

Vehicle:

unchanged (no vehicle)

Controls:

other: Not applicable

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied: Approximately 0.873 g of the solid test item was applied to the cornea. It covered adequately the corneal surface.

Duration of treatment / exposure:

240 minutes at 32±1°C

Number of animals or in vitro replicates:

Three corneas were allocated to the test item, three to the negative control and three to the positive control.

Details on study design:

Treatment of corneas
Approximately 0.873 g of the solid test item was found to adequately cover the corneal surface. 750 uL of each control item was applied to the appropriate corneas. The holders were gently tilted back and forth to ensure a uniform application of the item over the entire cornea. Each holder was incubated, anterior chamber uppermost, at 32± 1°C for 240 minutes. At the end of the exposure period the test item and control items were removed from the anterior chamber and the cornea was rinsed three times with fresh complete minimum essential medium (MEM) containing phenol red before a final rinse with complete MEM. the anterior and posterior chambers were refilled with fresh complete MEM. A post treatment opacity reading was taken and each cornea was visually observed.

Application of sodium fluorescein
Following the opacity measurement the permability of the corneas to sodium fluorescein was evaluated. The medium from the anterior chamber was removed and replaced with 1 mL of sodium fluorescein solution (5mg/mL). The dosing holes were plugged and the holders incubated, anterior chamber uppermost, at 32± 1°C for 90 minutes.

Permeability determinations
After incubation the medium in the posterior chamber of each holder was decanted and retained. 360 uL of medium representing each cornea was applied to a designated well on a 96-well plate and the optical density at 492 nm (OD492) was measured using the Anthos 2001 microplate reader.

Histopathology
The corneas were retained after testing for possible conduct of histopathology. Each cornea was placed into a pre-labelled tissue cassette fitted with a histology sponge to protect the endothelial surface. The cassette was immersed in 10% neutral buffered formalin.

Opacity measurement
The change in opacity for each cornea was calculated by subtracting the initial opacity reading from the final opacity reading. These values were then corrected by subtracting the average change in opacity observed for the negative control corneas. The mean opacity value of each treatment group was then calculated by averaging the corrected opacity values of each cornea for that treatment group.

Permeability measurement
The corrected OD492 was calculated by subtracting the mean OD492 of the negative control corneas from the OD492 value of each treated cornea. The OD492 value of each treatment group was calculated by averaging the corrected OD492 values of the treated corneas for the treatment group.

SCORING SYSTEM:
Results from the two test method endpoints, opacity and permeability, were combined in an empirically derived formula to generate an In Vitro Irritancy Score.
In Vitro Irritancy Score = mean opacity value + (15 x mean OD 492 value)

Additionally, the opacity and permeability values were evaluated independently to determine whether the test item induces a response through only one of the two endpoints.
A test item that induces an In Vitro Irritancy Score ≥55.1 is defined as an ocular corrosive or severe irritant and will be labelled EU DSD (67/548/EEC) R41 and EU CLP/UN GHS Category 1.

Irritation parameter:

in vitro irritation score

Run / experiment:

240 minutes

Value:

171.5

Vehicle controls validity:

valid

Positive controls validity:

valid

Remarks on result:

positive indication of irritation

Other effects / acceptance of results:

The test item was considered to be an ocular corrosive or severe irritant.

The corneas treated with the test item were opaque post incubation. The corneas treated with the negative control item were clear post incubation. The corneas treated with positive control item were cloudy post incubation.

The positive control In Vitro Irritancy Score was 82.1 and the negative control In Vitro Irritancy Score was 3.7.

Criterion for an Acceptable Test

The positive control In Vitro Irritancy Score was within the range of 65.9 to 140.8. The positive control acceptance criterion was therefore satisfied.

The In Vitro irritancy scores are summarized as follows:

 

Treatment	In Vitro Irritancy Score
Test Item	171.5
Negative Control	3.7
Positive Control	82.1

 

Table 1. Individual and Mean Corneal Opacity and Permeability Measurements

Treatment	Cornea Number	Opacity				Permeability (OD)		In VitroIrritancy Score
		Pre-Treatment	Post-Treatment	Post-Treatment-Pre‑Treatment	Corrected Value		Corrected Value
Negative Control	1	1	4	3		0.035
	2	1	4	3		0.067
	3	2	5	3		0.028
				3.0*		0.043¨		3.7
Positive Control	4	2	69	67	64.0	1.590	1.547
	5	2	58	56	53.0	0.759	0.716
	6	1	78	77	74.0	1.477	1.434
					63.7·		1.232·	82.1
Test Item	10	1	169	168	165.0	1.899	1.856
	11	2	144	142	139.0	2.360	2.317
	12	1	118	117	114.0	2.312	2.269
					139.3·		2.147·	171.5

OD= Optical density; * = Mean of the post-treatment -pre‑treatment values; ¨= Mean permeability; ·= Mean corrected value

Table 2. Corneal Epithelium Condition Post Treatment

 

Treatment	Cornea Number	Observation
		Post Treatment
Negative Control	1	clear
	2	clear
	3	clear
Positive Control	4	cloudy
	5	cloudy
	6	cloudy
Test Item	10	opaque
	11	opaque
	12	opaque

Interpretation of results:

Category 1 (irreversible effects on the eye) based on GHS criteria

Conclusions:

The test item was considered to be an ocular corrosive or severe irritant.

Executive summary:

Potassium dicyanoaurate was tested in vitro for its ocular irritancy potential to the isolated bovine cornea. The test item was applied neat for 240 minutes. Negative and positive control items were tested concurrently. The two endpoints, decreased light transmission through the cornea (opacity) and increased passage of sodium fluorescein dye through the cornea (permeability) were combined in an empirically derived formula to generate an In Vitro Irritancy Score (IVIS).

Potassium dicyanoaurate induces an In Vitro Irritancy Score of 171.5 and therefore was considered to be an ocular corrosive or severe irritant.

Reference 2

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10 March 1992

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliant, guideline test. Available as an unpublished report. Single animal tested for welfare reasons. Acceptable without restrictions.

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)

Deviations:

no

GLP compliance:

yes

Species:

rabbit

Strain:

other: White russian (Albino)

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: ASTA Medica AG. D-4802 Halle-Künsebeck
- Age at study initiation: 12 months
- Weight at study initiation: 2.42 kg
- Housing: Stainless steel cage with grating floor, type ASTA, size: 48.5x40x36.5
- Diet (e.g. ad libitum): ssniff K, Special diet for rabbits. Approx: 120 g/day.
- Water (e.g. ad libitum): ad libitum
- Acclimation period: One day

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.5 - 22.0°C
- Humidity (%): 50 - 55%
- Photoperiod (hrs dark / hrs light): 6 a.m. - 6 p.m. CET artificial lighting, 6 p.m. - 6 a.m. CET darkness.

Vehicle:

unchanged (no vehicle)

Controls:

not required

Amount / concentration applied:

0.101 g in one eye.

Duration of treatment / exposure:

Single application of test substacnce applied

Observation period (in vivo):

5 hours after treatment (animal was found dead at 7 hours after treatment)

Number of animals or in vitro replicates:

1

Details on study design:

REMOVAL OF TEST SUBSTANCE
None

SCORING SYSTEM:
DRAIZE scale as well as qualitative assessments.

TOOL USED TO ASSESS SCORE: Cliptrix pencil light

Irritation parameter:

cornea opacity score

Remarks:

opacity

Basis:

animal #1

Time point:

other: 1 hour after treatment

Score:

4

Max. score:

4

Reversibility:

not reversible

Irritation parameter:

cornea opacity score

Remarks:

area of cornea involved

Basis:

animal #1

Time point:

other: 1 hour after treatment

Score:

4

Max. score:

4

Reversibility:

not reversible

Irritation parameter:

iris score

Basis:

animal #1

Time point:

other: 1 hour after treatment

Score:

2

Max. score:

2

Reversibility:

not reversible

Irritation parameter:

conjunctivae score

Remarks:

redness

Basis:

animal #1

Time point:

other: 1 hour after treatment

Score:

3

Max. score:

3

Reversibility:

not reversible

Irritation parameter:

conjunctivae score

Remarks:

chemosis

Basis:

animal #1

Time point:

other: 1 hour after treatment

Score:

4

Max. score:

4

Reversibility:

not reversible

Irritation parameter:

conjunctivae score

Remarks:

discharge

Basis:

animal #1

Time point:

other: 1 hour after treatment

Score:

3

Max. score:

3

Reversibility:

not reversible

Irritant / corrosive response data:

All DRAIZE scores were at maximum 1 hour after treatment. No reversability occured - the animal was found dead 7 hours after treatment.
Severe hypokinesia and lacrimation at the non-treated eye 5 hours after treatment

Other effects:

Microscopial examination of the treated eye showed severe ulceration, moderate to severe coagulation necrosis and inflammatory cell infiltration in cornea and eyelids. Anterior eye chamber was filled with fibrinous masses and the iris exhibited moderate hypermia and severe edema.

Interpretation of results:

Category 1 (irreversible effects on the eye) based on GHS criteria

Conclusions:

Application of potassium dicyanoaurate to the eye of a rabbit caused severe changes 1 hour after treatment. The animal was found dead 7 hours after treatment.

Executive summary:

In a GLP, guideline test application of potassium dicyanoaurate to the eye of a rabbit caused severe changes 1 hour after treatment. The animal was found dead 7 hours after treatment. The test substance is therefore considered to be corrosive to the eye of the rabbit.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

The skin irritation potential of potassium dicyanoaurate was tested in vivo on rabbit skin. A solution of 0.5 g potassium dicyanoaurate in water was applied occlusively to three rabbits for four hours, followed by a 14-day observation period, according to OECD guideline 404 and EU method B.4 (Berthold 1992). Severe reactions were observed in all animals. Potassium dicyanoaurate was determined to be highly irritating. Skin irritation and corrosion were also tested with the EPISKIN reconstructed human epidermis model. Triplicate tissue samples were exposed to approximately 10 mg potassium dicyanoaurate applied topically for 15 minutes followed by a 42-hour post-exposure incubation period, according to OECD guideline 439 and EU method B.46 (Warren 2014a). Potassium dicyanoaurate is considered to be irritating based on EU criteria. In the skin corrosion test, duplicate tissue samples were exposed for 3, 60 and 240 minutes to approximately 20 mg potassium dicyanoaurate applied topically followed by a 3-hour post-exposure incubation period, according to OECD guideline 431 and EU method B.40 (Warren 2014b). Potassium dicyanoaurate is considered to be non-corrosive based on EU criteria.

Eye irritation was tested in vitro and in vivo. In the in vivo test performed according to OECD guideline 405 and EU method B.5, one rabbit was treated with a single application of 0.101 g potassium dicyanoaurate in one eye. All DRAIZE scores were at the maximum 1 hour after treatment and mortality occurred 7 hours after treatment. Severe hypokinesia and lacrimation at the non-treated eye were observed 5 hours after treatment. Potassium dicyanoaurate is considered to be corrosive to the rabbit eye (Berthold 1992). In the Bovine Corneal Opacity and Permeability (BCOP) assay following OECD guideline 437, eyes from adult cattle were treated with approximately 0.873 g of potassium dicyanoaurate for 240 minutes (Warren 2014c). Results of the opacity and permeability endpoints were combined to give an In Vitro Irritancy Score of 171.5. Potassium dicyanoaurate is considered to be an ocular corrosive or severe irritant based on EU criteria.

Overall, potassium dicyanoaurate is a skin irritant, an ocular corrosive and a severe eye irritant, but is not skin corrosive.

Justification for selection of skin irritation / corrosion endpoint:
Potassium dicyanoaurate was determined to be a highly irritating but non-corrosive to the skin in skin irritation and corrosion studies on rabbit skin and a human epidermis model. The in vivo rabbit study is considered to represent the most severe response.

Justification for selection of eye irritation endpoint:
Potassium dicyanoaurate was determined to be a severe eye irritant in the in vitro eye test and corrosive to the eye in the in vivo test. The in vivo rabbit study is considered to represent the most severe response.

Effects on skin irritation/corrosion: highly irritating

Effects on eye irritation: corrosive

Justification for classification or non-classification

According to GHS and EU CLP 1272/2008, potassium dicyanoaurate is classified as Category 2 for skin irritation, based on the results of in vivo rabbit and in vitro human epidermis model studies, and Category 1 (serious eye damage/irreversible effects on the eye) for effects on the eye, as serious eye damage was observed the in vivo eye irritation and BCOP studies.