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Toxicological information

Skin irritation / corrosion

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Administrative data

Endpoint:
skin irritation: in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Study start 20th September 2006 - Report date 16th February 2007
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2007
Report Date:
2007

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 404 (Acute Dermal Irritation / Corrosion)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.4 (Acute Toxicity: Dermal Irritation / Corrosion)
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid
Specific details on test material used for the study:
Identification: Witconate P-1460
Description: Light brown paste
Batch number: 2140-59.01
Activity: 88.8 %
Stability of test item: Stable under storage conditions.
Expiry date: 07-APR-2007
Storage conditions: At room temperature (range of 20 +/-5 °C), light protected

Test animals

Species:
rabbit
Strain:
New Zealand White
Details on test animals and environmental conditions:
Source of the rabbits: Harlan Netherlands BV Kreuzelweg 53, NL-5961 NM Horst / The Netherlands, Postbus 6174, NL-5960 AD Horst / The Netherlands
Number of animals per test: 3 (Animals of both sexes were used)
Age at treatment: 13 weeks (male), 10—11 weeks (females)
Identification: By unique cage number and corresponding ear number.
Acclimatization: Under laboratory conditions after health examination. Only animals without any visual signs of illness were used for the study.
Allocation: Male No. 85, Female Nos. 86, 87

HUSBANDRY
Conditions: Air-conditioned with ranges for room temperature 17-23 °C, relative humidity 30-70 % and approximately 10-15 air changes per hour. Room temperature and humidity were monitored continuously and values outside of these ranges may have occasionally occurred, usually following room cleaning. These transient variations are considered not to have any influence on the study and, therefore, these data are not reported but are retained at RCC. The animals were provided with an automatically controlled light cycle of 12 hours light and 12 hours dark. Music was played during the daytime light period.

Accomodation: Individually in stainless steel cages equipped with feed hoppers and drinking water bowls. Wood blocks (RCC Ltd, Füllinsdorf) and haysticks 4642 (batch no. 77/05, Provimi Kliba AG) were provided for gnawing.
Diet: Pelleted standard Provimi Kliba 3418 rabbit maintenance
diet ad Iibitum (batch no. 24/06) provided by Provimi Kliba AG, CH-4303 Kaiseraugst/Switzerland) ad Iibitum. Results of analyses for contaminants are archived at RCC Ltd.
Water: Community tap water from Füllinsdorf ad Iibitum. Results of bacteriological, chemical and contaminant analyses are archived at RCC Ltd.

Test system

Type of coverage:
semiocclusive
Preparation of test site:
clipped
Vehicle:
unchanged (no vehicle)
Amount / concentration applied:
0.5 g (per animal) of Witconate P-1460 was weighed as delivered by the Sponsor.
The pH of the test item was measured before the study initiation date. A formulation of a 1 % (w/w) solution was prepared. The pH was found to be 5-6.
According to Commission Directive 2004/73/EC, 8.4. and OECD Guidelines 404, a test item needs not to be tested if the pH-value is less than 2 or greater than 11.5, owing to its predictable corrosive properties.
Duration of treatment / exposure:
Four days before treatment, the left flank was clipped with an electric clipper, exposing an area of approximately 100 cm2 (10 cm x 10 cm). The skin of the animals was examined one day before treatment. and regrown fur of all animals was clipped again.
Animals with overt signs of skin injury or marked irritation which may have interfered with the interpretation of the results were not used in the test.
On the day of treatment, 0.5 g of Witconate P-1460 was placed on a surgical gauze patch (ca. 4 cm x 4 cm). This gauze patch was applied to the intact skin of the clipped area. The patch was covered with a semi-occlusive dressing. The dressing was wrapped around the abdomen and anchored with tape.
The duration of treatment was 4 hours. Then the dressing was removed and the skin was flushed with lukewarm tap water to clean the application site so that any reactions (erythema) were clearly visible at that time.
As it was suspected that the test item might produce irritancy, a single animal (one female) was treated first. Because moderate signs of irritation were observed following the treatment. the two remaining animals were also treated in a stepwise manner.
Observation period:
Mortality/Viability: Daily from acclimatization of the animals to the termination of test.
Body weights: At start of acclimatization, on the day of application and at termination of observation.
Clinical signs: Daily from acclimatization of the animals to the termination of test.
Skin reactions: where recorded
Number of animals:
3 (Animals of both sexes were used), Male No. 85, Female Nos. 86, 87
Details on study design:
IRRITATION SCORES
The skin reaction was assessed according to the numerical scoring system listed in the Commission Directive 2004/73/EC, April 29, 2004.
If evident, corrosive or staining properties of the test item were described and recorded.
SKIN OBSERVATION TIME POINTS AND TERMINATION
The skin reaction was assessed at approximately 1, 24, 48 and 72 hours, as well as 7, 10 and 14 days after exposure (removal of the dressing, gauze patch and test item).
To allow further examination of the test site, animal No. 85 was re-clipped on completion of the 48 hour examination.

TREATMENT OF RESULTS
Data were summarized in tabular form, showing for each individual animal the irritation scores for erythema and oedema at all measurement intervals. Any lesions were described, including the degree and nature of irritation, corrosion or any other toxic effects observed, and their reversibility.
The mean score was calculated across 3 scoring times (24, 48 and 72 hours after patch removal) for each animal for erythema/eschar grades and for oedema grades, separately. An animal is positive when the mean score is 2 or greater. The test is positive for irritation when at least 2 animals are positive for the same endpoint (erythema/eschar or oedema).

NECROPSY
No necropsy was performed on the animals sacrificed at termination of observation. All rabbits were sacrificed by an intravenous injection of pentobarbitone into the ear vein at a dose of at least 1 mL/kg body weight and discarded.

Results and discussion

In vivo

Resultsopen allclose all
Irritation parameter:
erythema score
Basis:
animal #1
Time point:
24/48/72 h
Score:
2
Max. score:
2
Reversibility:
fully reversible within: 10 days
Remarks on result:
probability of moderate irritation
Irritation parameter:
edema score
Basis:
animal #1
Time point:
24/48/72 h
Score:
1
Max. score:
2
Reversibility:
fully reversible within: 7 days
Remarks on result:
probability of mild irritation
Irritation parameter:
erythema score
Basis:
animal #2
Time point:
24/48/72 h
Score:
2
Max. score:
2
Reversibility:
fully reversible within: 7 days
Remarks on result:
probability of moderate irritation
Irritation parameter:
edema score
Basis:
animal #2
Time point:
24/48/72 h
Score:
1.33
Max. score:
2
Reversibility:
fully reversible within: 7 days
Remarks on result:
probability of moderate irritation
Irritation parameter:
erythema score
Basis:
animal #3
Time point:
24/48/72 h
Score:
1
Max. score:
2
Reversibility:
fully reversible within: 7 days
Remarks on result:
probability of mild irritation
Irritation parameter:
edema score
Basis:
animal #3
Time point:
24/48/72 h
Score:
0.33
Max. score:
2
Reversibility:
fully reversible within: 7 days
Remarks on result:
probability of weak irritation
Irritant / corrosive response data:
IRRITATION
The mean score was calculated across 3 scoring times (24, 48 and 72 hours after patch removal) for each animal for erythema/eschar grades and for oedema grades, separately. The mean erythema/eschar score of the three animals was 2.00, 2.00 and 1.00, respectively and the mean oedema score was 1.00, 1.33 and 0.33, respectively.
A well-defined erythema was observed at the treatment site from the 1-hour reading up to the 24- or 72-hour reading in one and two animals, respectively. A very slight erythema was noted in one animal 24 hours after the treatment and in one animal still at the 7-day reading.
A slight swelling was noted for the three animals one hour after end of the exposure and persisted in one animal still at the 24-hour reading. A very slight swelling was observed 24 to 72 or 48 to 72 hours after the treatment, or, at the 24-hour reading only.
Scaling of the skin at the application site was noted in two of the animals 72 hours to 7 days or 7 to 14 days after the treatment, respectively.

COLORATION

No staining produced by the test item of the treated skin was observed.

CORROSION

Neither alterations of the treated skin were observed nor were corrosive effects evident on the skin.
Other effects:
VIABILITYIMORTALITYICLINICAL SIGNS

No clinical signs of systemic toxicity were observed in the animals during the study and no mortality occurred.


The body weights of all rabbits were considered to be within the normal range of variability.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
Based upon the referred classification criteria (Commission Directive 2001/59/EC of August 2001), Witconate P-1460 was considered to be "Irritating” to rabbit skin and needed to be labelled with the risk phrase R38 “Irritating to skin”. The classification was based on the EU criteria as two out of the three rabbits had a mean erythema score for 24/48/72 hours of 2. The corresponding EU CLP/GHS criteria for skin irritation requires for the equivalent classification of category 2, a mean value in at least two of the three rabbits over 24/48/72 hours for erythema and or oedema of >2.3-< 4.0. Due to this change in the threshold from 2.0 used in the previous EU criteria for R38 to 2.3 for category 2, Witconate P-1460 does not meet the CLP/GHS criteria to be classified as irritating to skin. It also does not meet the criteria as the effects (erythema and or oedema) did not persist to 14 days with the exception of scaling in one rabbit. For scaling to result in a classification of category 2 for skin irritation it must persist for 14 days in at least 2 of the three rabbits.
Executive summary:

The primary skin irritation potential of Witconate P-1460 was investigated according to OECD test guideline no. 404. The test item was applied by topical semi-occlusive application of 0.5 g to the intact left flank of each of three young adult New Zealand White rabbits. The duration of treatment was four hours. The scoring of skin reactions was performed 1, 24, 48 and 72 hours, as well as 7, 10 and 14 days after removal of the dressing.

The mean score was calculated across 3 scoring times (24, 48 and 72 hours after patch removal) for each animal for erythema/eschar grades and for oedema grades, separately.

The mean erythema/eschar score of the three animals was 2.00, 2.00 and 1.00, respectively and the mean oedema score was 1.00, 1.33 and 0.33, respectively.

The application of Witconate P-1460 to the skin resulted in moderate signs of irritation.

These included erythema, oedema and scaling. These effects were not fully reversible in one of the treated animals during the 14-day observation period. The test item caused no staining of the treated skin. No corrosive effects were noted on the treated skin of any animal at any of the measuring intervals and no clinical signs were observed.

Based upon the referred classification criteria (Commission Directive 2001/59/EC of August 2001), Witconate P-1460 is considered to be “Irritating" to rabbit skin and needs to be labelled with the risk phrase R38 “Irritating to skin".

The classification was based on the EU criteria as two out of the three rabbits had a mean erythema score for 24/48/72 hours of 2. The corresponding EU CLP/GHS criteria for skin irritation requires for the equivalent classification of category 2, a mean value in at least two of the three rabbits over 24/48/72 hours for erythema and or oedema of >2.3-<4.0. Due to this change in the threshold from 2.0 used in the previous EU criteria for R38 to 2.3 for category 2, Witconate P-1460 does not meet the CLP/GHS criteria to be classified as irritating to skin. It also does not meet the criteria as the effects (erythema and or oedema) did not persist to 14 days with the exception of scaling in one rabbit. For scaling to result in a classification of category 2 for skin irritation it must persist for 14 days in at least 2 of the three rabbits.