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Description of key information

In two in vitro EpiSkin assays conducted in accordance with OECD Test Guidelines 431 and 439 (corrosion and irritation, respectively) and to GLP, dihydrogen hexahydroxyplatinate was considered to be non-corrosive and non-irritating to skin (Kiss, 2012a, b).

 

In an OECD Test Guideline 405 in vivo eye irritation study, conducted to GLP, hexahydroxoplatinum(IV) acid produced some transient irritation following instillation of the test material (0.1 ml) into one eye of each of 3 rabbits (Berthold, 1995b).

 

No relevant respiratory tract irritation data were identified.

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:
22-24 December 2011
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Good-quality, well-documented guideline study, to GLP
Qualifier:
according to
Guideline:
other: OECD Guidelines for Testing of Chemicals, Section 4, No. 439, “In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method” adopted 22 July 2010
Deviations:
no
Qualifier:
according to
Guideline:
other: EpiSkin™ SOP, Version 1.8 (February 2009), ECVAM Skin Irritation Validation Study: Validation of the EpiSkin™ test method 15 min - 42 hours for the prediction of acute skin irritation of chemicals. Available at: [http://ecvam.jrc.ec.europa.eu]
Deviations:
no
Qualifier:
according to
Guideline:
other: IN VITRO SKIN IRRITATION: RECONSTRUCTED HUMAN EPIDERMIS MODEL TEST in relation to Regulation (EC) No 440/2008 (as amended) and Regulation (EC) No 1907/2006 on REACH (Annex III, B.46).
Deviations:
no
Principles of method if other than guideline:
The test is designed to predict and classify the skin irritant potential of chemicals according to chemical safety regulations, using the reconstructed human epidermis model EPISKIN-SM and parameters related to skin irritation.
EPISKIN-SM is a three-dimensional human skin model comprising a reconstructed epidermis with a functional stratum corneum. Its use for skin irritation testing involves topical application of test materials to the surface of the epidermis, and the subsequent assessment of their effects on cell viability. Cell viability determination is based on cellular mitochondrial dehydrogenase activity, measured by MTT reduction and conversion into a blue formazan salt that is quantitatively measured after extraction from tissues. The reduction of cell viability in treated tissues is compared to negative controls and expressed as a %. The % reduction in viability is used to predict the irritation potential.

The EPISKIN-SM has been found scientifically valid for reliably predicting no label and R38 (irritant) substances in respect to the previous EU classification scheme and has been confirmed in April 2009 by ESAC for use under the UN GHS system as "applicable to all authorities". It is approved by international regulatory agencies as a replacement for the identification of irritants/corrosives in the in vivo rabbit skin assay(OECD 404).
GLP compliance:
yes (incl. certificate)
Species:
human
Strain:
other: Reconstructed human epidermis model (see details below)
Details on test animals and environmental conditions:
EPISKIN-SM (Source: SkinEthic, France, Batch No.:11-EKIN-047, Expiry date: 26 December 2011) is a three-dimensional human epidermis model. Adult human-derived epidermal keratinocytes are seeded on a dermal substitute consisting of a collagen type I matrix coated with type IV collagen. A highly differentiated and stratified epidermis model is obtained after 13-day culture period comprising the main basal, supra basal, spinous and granular layers and a functional stratum corneum.
Type of coverage:
other: Applied evenly to the epidermal surface following the application of 10 ul distilled water to this surface
Preparation of test site:
other: in vitro cell culture
Vehicle:
water
Remarks:
distilled
Controls:
other: negative control skin unit tested in triplicate
Amount / concentration applied:
20 mg (finely ground) to each of three test skin units.
10 µl PBS (phosphate buffered saline) was added to each of the three negative control skin units and 10 µl SDS (sodium dodecyl sulfate, 5% aqueous solution) was added to each of the three positive control skin units.
For additional control for staining effects of the test item, 10 µl distilled water was applied to the epidermal surface of a single skin unit to ensure good contact with the epidermis, then 20 mg of the test item (finely ground) was applied evenly to the epidermal surface.
Duration of treatment / exposure:
Exposure for 15 minutes (± 0.5 min) at room temperature (20-37°C).
Then incubated with fresh “maintenance medium” for 42 hours (± 1h) at 37°C.
Observation period:
Not applicable to this test system
Number of animals:
Not applicable to this test system
Details on study design:
EPISKIN-SM assay plate contained reconstructed epidermis units (area: 0.38 cm2); each was attached to the base of a tissue culture vessel and maintained on nutritive agar.

After test substance exposure and subsequent incubation, preparations for cell viability determination were: incubation with MTT solution (at 37 degrees C for 3 hours) followed by incubation with acidified isopropanol for formazan extraction (around two hours at room temperature with gentle agitation).

For cell viability measurements, the OD (Absorbance / Optical Density) of the samples in a spectrophotometer was read at 540 nm using acidified isopropanol solution blank (6×200 µL). (The validity of the microplate reader was verified with a standard verification plate daily before use. The standard plate was calibrated yearly by the manufacturer.)

For each treated tissue, OD (as adjusted for colouring potential of the test substance) was calculated and the tissue viability was expressed as a % relative to negative control.

Criteria for classification as irritant/non-irritant: If the resulting mean relative viability (as adjusted for intrinsic colour) is less than or equal to 50% of the negative control, the test substance is considered to be irritant to skin.

Irritation / corrosion parameter:
other: other:
Value:
82
Remarks on result:
other:
Remarks:
Basis: mean Cell/tissue viability. Time point: 42 hours. Reversibility: other: Not applicable. Remarks: Score is a percentage (%) of negative control. Mean relative viability <=50% the test substance is considered to be irritant to skin.. (migrated information)
Other effects / acceptance of results:
Mean cell viability (as adjusted for intrinsic colour) was 82% of the negative control (range 74%-88%).
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
In an in vitro EpiSkin assay conducted in accordance with OECD Test Guideline 439 and to GLP, dihydrogen hexahydroxyplatinate was considered to be non-irritating to skin.
Executive summary:

Dihydrogen hexahydroxyplatinate was tested for skin irritation potential in an in vitro reconstructed human epidermis model (EpiSkin assay) conducted in accordance with OECD Test Guideline 439, and to GLP.

 

EpiSkin is a three-dimensional human skin model comprising a reconstructed epidermis with a functional stratum corneum. Its use for skin irritation testing involves topical application of test materials to the surface of the epidermis, and the subsequent assessment of their effects on cell viability. Cell viability determination is based on cellular mitochondrial dehydrogenase activity, measured by MTT reduction and conversion into a blue formazan salt that is quantitatively measured after extraction from tissues. The reduction of cell viability in treated tissues is compared to negative controls and expressed as a %. If the resulting mean relative viability (as adjusted for intrinsic colour) is less than or equal to 50% of the negative control, the test substance is considered to be irritant to skin.

 

Following a 15-minute exposure to the test substance, the test system skin cell viability was calculated to be greater than 50% (the average was 82% and the range was 74 -88%), and it was therefore considered to be non-irritating to skin.

 

Under the conditions of this assay, dihydrogen hexahydroxyplatinate does not require classification for skin irritation according to EU CLP criteria (EC 1272/2008).

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:
20 March - 31 March 1995
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Guideline study, to GLP, with minor deviations (humidity) not anticipated to affect the valididty of the study.
Qualifier:
according to
Guideline:
OECD Guideline 405 (Acute Eye Irritation / Corrosion)
Deviations:
yes
Remarks:
The relative humidity was outside the stated range on 2 days; this was considered to have no influence on the results of the study
Qualifier:
according to
Guideline:
other: EEC Guideline 92/32/EEC
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-33790 Halle/Westfalen
- Age at study initiation: 33-49 months
- Weight at study initiation: 2.22-2.94 kg
- Housing: Stainless steel cages with grating floor, type ASTA, size: 48.5x40x36.5 (LxBxH); 1 animal per cage
- Diet (e.g. ad libitum): Approximately 120 g/day/animal of Standard diet, ssniff K, Special diet for rabbits
- Water (e.g. ad libitum): ad libitum from municipal supply
- Acclimation period: at least one day

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.5 degrees centigrade "constantly"
- Humidity (%): 32-72%
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12/12
Vehicle:
unchanged (no vehicle)
Controls:
not required
Amount / concentration applied:
63.4-85.1 mg (representing a volume of around 0.1 ml) of the test substance was applied into the conjunctival sac of one eye; both lids were briefly closed by gentle finger pressure. Eyes were not rinsed.
Duration of treatment / exposure:
Single application; no rinse was applied during observation period of up to 4 days.
Observation period (in vivo):
3 days for 2 animals; 4 days for one animal.
Observation at 1, 24, 48 and 72 hours after a single application. One animal was observed after an additional 24 hours.
Number of animals or in vitro replicates:
3 males
Details on study design:
SCORING SYSTEM: Quantitative and qualitative assessment using the Draize scale (See "Any other information on materials and methods ...", below). An irritation index was calculated from these scores and used to grade irritant effects using a modified method according to Gilman et al.

TOOL USED TO ASSESS SCORE:Cliptrix pencil light; visual assessment, presumably by an experienced technician.

Eyes were examined for corrosive effects; animals were assessed for clinical symptoms that may indicate systemic toxic effects.
Irritation parameter:
other: Irritation index
Basis:
mean
Time point:
other: 1, 24, 48 and 72 hours post-application
Score:
6
Max. score:
110
Reversibility:
other: Not applicable
Irritant / corrosive response data:
Some degree of conjunctival redness was seen in all animals. In two, this was slight or moderate and was fully reversible within 3 days; in the remaining animal, moderate to severe redness was reported over 3 days, but was not present by day 4.

Severe conjunctival swelling (chemosis; Draize score 3 of a maximum of 4) was seen in one animal one day following instillation of the test substance; this decreased gradually and was not present by day 4. No conjunctival swelling was seen in the remaining two animals throughout the observation period.

Severe conjunctival discharge (maximum Draize score of 3) was seen in one animal immediately after the test substance was applied, but cleared within 24 hours. In another, abnormal discharge was reported over 2 days following application of the test substance (at 24 hours, this was severe, with the maximum Draize score of 3), but had cleared by day 3. One animal exhibited no abnormal discharge throughout the observation period.

Effects on the iris were seen in one animal at 1 hour (slight circumcorneal hyperemia), and in another animal at 24 and 48 hours (moderate circumcorneal hyperemia). All effects were classified as grade 1 on the Draize scale (where there is a maximum score of 2). In both cases the iris was normal at 3 days. In one animal, the iris was normal throughout the observation period.

No corneal effecs (opacity) were observed in any animal.

No systemic toxic effects were observed.

Draize scoring for ocular lesions:

Cornea

 

a. Opacity-degree of density (Area most dense taken for reading)

 

    No ulceration or opacity                                                                                   0

    Scattered or diffuse areas of opacity (other than slight

    dulling or normal lustre): details of iris clearly visible                                      1

    Easily discernible translucent area: details of iris slightly obscured                 2

    Nacreous area: no details of iris visible: size of pupil barely discernible          3

    Opaque cornea: iris not discernible through the opacity                                   4

 

 

b.  Area of cornea involved

 

One quarter (or less), but not zero                                                                    1

Greater than one quarter, but less than half                                                      2

Greater than half, but less than three quarters                                                  3

Greater than three quarters, up to whole area                                                   4

 

 

Iris

 

a. Values

 

Normal                                                                                                              0

Markedly deepened rugae, congestion, swelling, moderate

circumcorneal hyperaemia: or injection: iris reactive to light

(a sluggish reaction is considered to be an effect)                                           1

Haemorrhage, gross destruction, or no reaction to light.                                 2

Conjunctivae

 

a. Redness (Palpebral and bulbar)

 

Normal                                                                                                              0

Some blood vessels hyperaemic (injected)                                                       1

Diffuse, crimson colour, individual vessels not easily discernible                   2

Diffuse beefy red                                                                                             3

 

 

b.Chemosis

 

Normal                                                                                                              0

Some swelling above normal (includes nictating membrane)                           1

Obvious swelling with partial eversion of lids                                                 2

Swelling with lids about half closed                                                                3

Swelling with lids more than half closed                                                         4

 

 

c. Discharge

 

No discharge                                                                                                    0

Any amount different from normal (does not include small amounts

observed in inner canthus of normal animals)                                                  1

Discharge with moistening of the lids and hairs just adjacent to lids              2

Discharge with moistening of the lids and hairs, on considerable area

around the eye                                                                                                  3

 

Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
In an OECD Test Guideline 405 in vivo eye irritation study, conducted to GLP, hexahydroxoplatinum(IV) acid produced some transient irritation following instillation of the test material (0.1 ml) into one eye of each of 3 rabbits.
Executive summary:

In an in vivo eye irritation study carried out in accordance with OECD Test Guideline 405, and to GLP, a 0.1 ml aliquot of undiluted hexahydroxoplatinum(IV) acid was instilled into the conjunctival sac of each of three male White Russian rabbits. The other eye was untreated to serve as a control. Following instillation the eyelids were held closed by gentle finger pressure. The ocular response was assessed at 1, 24, 48 and 72 hours. One animal was observed after an additional 24 hours. Corneal opacity, effects on the iris and conjunctival redness, chemosis and discharge were scored according to the Draize numerical scale.

 

No corneal effects (opacity) were observed in any animal. Effects on the iris were seen in one animal at 1 hour (slight circumcorneal hyperemia), and in another animal at 24 and 48 hours (moderate circumcorneal hyperemia). Effects on the conjunctiva were seen in all animals. Notably, in one animal, these were moderate/severe, with maximum scores for discharge and redness reported for at least one time point within the observation period. Indeed, the maximum observed score for conjunctival chemosis (swelling) in this animal was 3 (of a maximum of 4 achievable on the Draize scale). All effects on the iris and conjunctivae were, however, fully reversible within 4 days.

 

The primary irritation index calculated for all animals was 6 (out of 110). The study authors considered the substance to be non-irritant to rabbit eyes. Based on the severe irritation observed in one of the tested animals, it is prudent to classify the test item as an eye irritant (category 2), according to EU CLP criteria (EC 1272/2008).

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

No relevant human irritation/corrosion data were identified. No in vitro eye irritation studies were identified, or are required, as a reliable in vivo study is already available.

 

Dihydrogen hexahydroxyplatinate was tested for skin corrosion potential in an in vitro reconstructed human epidermis model (EpiSkin assay) conducted in accordance with OECD Test Guideline 431, and to GLP. EpiSkin is a three-dimensional human skin model comprising a reconstructed epidermis with a functional stratum corneum. Its use for skin corrosivity testing involves the topical application of test materials to the surface of the skin, and the subsequent assessment of their effects on cell viability. Cytotoxicity is expressed as the reduction of mitochondrial dehydrogenase activity measured by formazan production from MTT. The reduction of cell viability in treated tissues is compared to negative controls and expressed as a %. If the resulting mean relative viability is less than 35% of the negative control, the test substance is considered to be corrosive to skin. Following a 4-hr incubation with the test substance, the test system skin cell viability was calculated to be greater than 35% (the average was 82% and the range was 72 -89%), and it was therefore considered to be non-corrosive to skin (Kiss, 2012a). Under the conditions of this assay, dihydrogen hexahydroxyplatinate does not require classification for skin corrosion according to EU CLP criteria (EC 1272/2008).

 

Subsequently, dihydrogen hexahydroxyplatinate was tested for skin irritation potential in a further EpiSkin assay conducted in accordance with OECD Test Guideline 439, and to GLP. The assay designed to assess skin irritation is highly similar to the analogous assay for corrosivity. In this assay, if the mean relative viability (as adjusted for intrinsic colour) is less than or equal to 50% of the negative control, the test substance is considered to be irritant to skin. Following a 15-minute exposure to the test substance, the test system skin cell viability was calculated to be greater than 50% (the average was 82% and the range was 74 -88%), and it was therefore considered to be non-irritating to skin (Kiss, 2012b). Under the conditions of this assay, dihydrogen hexahydroxyplatinate does not require classification for skin irritation according to EU CLP criteria (EC 1272/2008).

 

In an in vivo eye irritation study carried out in accordance with OECD Test Guideline 405, and to GLP, a 0.1 ml aliquot of undiluted hexahydroxoplatinum(IV) acid was instilled into the conjunctival sac of each of three male White Russian rabbits. The other eye was untreated to serve as a control. Following instillation the eyelids were held closed by gentle finger pressure. The ocular response was assessed at 1, 24, 48 and 72 hours. One animal was observed after an additional 24 hours. Corneal opacity, effects on the iris and conjunctival redness, chemosis and discharge were scored according to the Draize numerical scale. No corneal effects (opacity) were observed in any animal. Effects on the iris were seen in one animal at 1 hour (slight circumcorneal hyperemia), and in another animal at 24 and 48 hours (moderate circumcorneal hyperemia). Effects on the conjunctiva were seen in all animals. Notably, in one animal, these were moderate/severe, with maximum scores for discharge and redness reported for at least one time point within the observation period. Indeed, the maximum observed score for conjunctival chemosis (swelling) in this animal was 3 (of a maximum of 4 achievable on the Draize scale). All effects on the iris and conjunctivae were, however, fully reversible within 4 days. The primary irritation index calculated for all animals was 6 (out of 110). The study authors considered the substance to be non-irritant to rabbit eyes (Berthold, 1995b). Based on the severe irritation observed in one of the tested animals, it is prudent to classify the test item as an eye irritant (category 2), according to EU CLP criteria (EC 1272/2008).

 

No respiratory tract irritation data were identified. A new study was not conducted as it is not a REACH Standard Information Requirement.

Justification for classification or non-classification

Based on the results of the available reliable in vitro skin irritation study, there is no requirement to classify dihydrogen hexahydroxyplatinate for skin irritation under EU CLP criteria (EC 1272/2008).

 

Based on the results of the available and reliable in vivo eye irritation study (severe irritation observed in one of the tested animals), it is prudent to classify the test item as an eye irritant (category 2), according to EU CLP criteria (EC 1272/2008).