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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:
Between 31 August 2011 and 04 October 2011
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. The study report was conclusive, done to a valid guideline and the study was conducted under GLP conditions.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2011
Report Date:
2011

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
Type:
Constituent
Type:
Constituent
Details on test material:
Sponsor's identification: Benzenesulfonic acid, 4-C10-13-sec-alkyl derivs., compds. with N-[3-(dimethylamino)propyl]-2-methyl-2-propenamide (1:1)
Description : light amber coloured viscous liquid
Batch number: P1113
Purity : 96.3 w%
Date received: 01 July 2011
Expiry date : 30 March 2012
Storage conditions: room temperature in the dark

The integrity of supplied data relating to the identity, purity and stability of the test item is the responsibility of the Sponsor.

A Certificate of Analysis supplied by the Sponsor is given in Appendix 1 - (attachment 1)

Test animals

Species:
rabbit
Strain:
New Zealand White
Details on test animals and environmental conditions:
Two New Zealand White (Hsdlf:NZW) strain rabbits were used.
At the start of the study the animals weighed 2.28 or 2.55 kg and were twelve to twenty weeks old. After an acclimatisation period of at least five days each animal was given a number unique within the study which was written with a black indelible marker-pen on the inner surface of the ear and on the cage label.

The animals were individually housed in suspended cages. Free access to mains drinking water and food was allowed throughout the study. The diet and drinking water were considered not to contain any contaminant of a level that might have affected the purpose or integrity of the study.
The temperature and relative humidity were set to achieve limits of 17 to 23°C and 30 to 70% respectively. Any occasional deviations from these targets were considered not to have affected the purpose or integrity of the study. The rate of air exchange was at least fifteen changes per hour and the lighting was controlled by a time switch to give twelve hours continuous light (06:00 to 18:00) and twelve hours darkness.
The animals were provided with environmental enrichment items which were considered not to contain any contaminant of a level that might have affected the purpose or integrity of the study.

Test system

Type of coverage:
semiocclusive
Preparation of test site:
other: clipped
Vehicle:
unchanged (no vehicle)
Controls:
no
Amount / concentration applied:
A quantity of 0.5 ml of the test item was applied directly to the skin under a 2.5 cm x 2.5 cm cotton gauze patch.
Duration of treatment / exposure:
Up to 4 hours
Observation period:
Up to 14 days
Number of animals:
2
Details on study design:
On the day before the test each rabbit was clipped free of fur from the dorsal/flank area using veterinary clippers. Only animals with a healthy intact epidermis by gross observation were selected for the study.
One rabbit was initially treated. Three suitable sites were selected on the back of the rabbit. A quantity of 0.5 ml of the test item was applied directly to the skin under a 2.5 cm x 2.5 cm cotton gauze patch. Each patch was secured in position with a strip of surgical adhesive tape. To prevent the animal interfering with the patches, the trunk of the rabbit was wrapped in an elasticated corset for the duration of the exposure period.
One patch was removed at each of three time points: 3 minutes, 1 hour and 4 hours after application. Any residual test item was removed by gentle swabbing with cotton wool soaked in 74% Industrial Methylated Spirits.
After consideration of the skin reactions produced in the first animal, an additional animal was treated with 0.5 ml of test item. One patch was applied to the back of the rabbit and was allowed to remain in contact with the skin for a period of four hours.

Results and discussion

In vivo

Resultsopen allclose all
Irritation parameter:
other: Erythema/Eschar Formation
Basis:
animal: 70905 Male
Time point:
other: Score at 1, 24, 48 and 72 hours, 7 and 14 days
Score:
2
Max. score:
4
Reversibility:
not fully reversible within: 14 days
Remarks on result:
other: Loss of skin elasticity and flexibility, crust formation and glossy skin noted. Adverse reaction prevented accurate evaluation of erythema at 7 days.
Irritation parameter:
other: Erythema/Eschar Formation
Basis:
animal: 709830 Male
Time point:
other: Score at 1, 24, 48 and 72 hours, 7 and 14 days
Score:
2
Max. score:
4
Reversibility:
not fully reversible within: 14 days
Remarks on result:
other: Loss of skin elasticity and flexibility, crust formation, light brown discoloration of the epidermis, hardened light brown coloured scab and glossy skin noted. Adverse reaction prevented accurate evaluation of erythema at 7 days.
Irritation parameter:
other: Oedema Formation
Basis:
animal: 70905 Male
Time point:
other: Score at 1, 24, 48 and 72 hours, 7 and 14 days
Score:
2
Max. score:
4
Reversibility:
fully reversible within: 7 days
Irritation parameter:
other: Oedema Formation
Basis:
animal: 70983 Male
Time point:
other: Score at 1, 24, 48 and 72 hours, 7 and 14 days
Score:
2
Max. score:
4
Reversibility:
fully reversible within: 14 days
Remarks on result:
other: Adverse reaction prevented accurate evaluation of oedema at 7 days.
Irritant / corrosive response data:
3-Minute Exposure Period
The individual scores for erythema/eschar and oedema are given in Table 1.
Well-defined erythema and very slight oedema were noted at the treated skin site at the 24, 48 and 72 Hour observations. Crust formation, preventing accurate evaluation of erythema, was noted at the treated skin site at the 7 Day observation. Glossy skin was noted at the treated skin site at the 14 Day observation.

1-Hour Exposure Period
The individual scores for erythema/eschar and oedema are given in Table 1.
Well-defined erythema and slight oedema were noted at the treated skin site at the 24, 48 and 72 Hour observations. Crust formation, preventing accurate evaluation of erythema, was noted at the treated skin site at the 7 Day observation. Glossy skin was noted at the treated skin site at the 14 Day observation.

4-Hour Exposure Period
The individual scores for erythema/eschar and oedema are given in Table 2.
Well-defined erythema and slight oedema were noted at one treated skin site immediately after patch removal. Well-defined erythema and slight oedema were noted at one treated skin site with very slight erythema and slight oedema noted at the other treated skin site one hour after patch removal. Well defined erythema and slight oedema were noted at both treated skin sites at the 24, 48 and 72 Hour observations.
Loss of skin flexibility and/or elasticity was noted at both treated skin sites at the 24, 48 and 72 Hour observations. Light brown discolouration of the epidermis was also noted at one treated skin site at the 24, 48 and 72-Hour observations. Crust formation was noted at one treated skin site at the 72 Hour observation and at the other treated skin site at the 7 Day observation. Hardened light brown coloured scab was noted at one treated skin site at the 7-Day observation. Adverse reactions prevented accurate evaluation of oedema and/or erythema at both treated skin sites at the 7 Day observation. Glossy skin was noted at both treated skin sites at the 14 Day observation.
Other effects:
Bodyweight

Individual bodyweights and bodyweight changes are given in Table 3.
Both animals showed expected gain in bodyweight during the study.

Any other information on results incl. tables

Interpretation of Results

Calculation of Primary Irritation Index and Grading of Irritancy Potential Using the Draize Scheme

The scores for erythema and oedema at the 24 and 72‑Hour readings were totalled for the two test rabbits (8 values) and this total was divided by four to give the primary irritation index of the test item. The test item was classified according to the following scheme devised by Draize J H (1959) "Dermal Toxicity" In: Appraisal of the Safety of Chemicals in Foods, Drugs and Cosmetics. Association of Food and Drug Officials of theUnited States,Austin,Texas, p.46‑59:

Primary Irritation Index

Classification of Irritancy

     0

Non-irritant

> 0 to 2

Mild irritant

> 2 to 5

Moderate irritant

> 5 to 8

Severe irritant

If irreversible alteration of the dermal tissue is noted in any rabbit, as judged by the Study Director, which include ulceration and clear necrosis or signs of scar tissue, the test item is classified as corrosive to rabbit skin. Classification according to Draize may, therefore, not be applicable.

The results were also interpreted according to EU labelling regulations Commission Directive 2001/59/EC and Regulation (EC) No 1272/2008 for the classification, packaging and labelling of dangerous substances.

Table 1              Individual Skin Reactions Following 3-Minute and 1-Hour Exposures

Skin Reaction

Observation Time
(following patch removal)

Individual Scores - Rabbit Number and Sex

70905Male

3-Minute Exposure

1-Hour Exposure

Erythema/Eschar Formation

Immediately

0

0

1 Hour

0

0

24 Hours

2

2

48 Hours

2

2

72 Hours

2

2

7 Days

?eCf

?eCf

14 Days

0G

0G

Oedema Formation

Immediately

0

0

1 Hour

0

0

24 Hours

1

2

48 Hours

1

2

72 Hours

1

2

7 Days

0

0

14 Days

0

0


Cf =     Crust formation

G =      Glossy skin

?e =     Adverse reaction prevented accurate evaluation of erythema

Table 2              Individual Skin Reactions Following 4-Hour Exposure

Skin Reaction

Observation Time
(following patch removal)

Individual Scores – Rabbit Number and Sex

Total

70905Male

70983Male

Erythema/Eschar Formation

Immediately

0

2

(2 )

1 Hour

1

2

( 3 )

24 Hours

2Le

2BrLeLf

4

48 Hours

2LeLf

2BrLeLf

( 4 )

72 Hours

2Le

2BrLeLfCf

4

7 Days

?eCf

?eSp

( 0-8 )

14 Days

0G

0G

( 0 )

Oedema Formation

Immediately

0

2

( 2 )

1 Hour

2

2

( 4 )

24 Hours

2

2

4

48 Hours

2

2

( 4 )

72 Hours

2

2

4

7 Days

0

?od

( 0-4 )

14 Days

0

0

( 0 )

Sum of 24 and 72-hour Readings (S)       :          16

Primary Irritation Index (S/4)                   :          16/4 = 4.0

Classification                                        :          MODERATE IRRITANT


(   ) = Total values not used for calculation of primary irritation index

Le =     Loss of skin elasticity

Lf =      Loss of skin flexibility

Br =     Light brown discolouration of the epidermis

Cf =     Crust formation

Sp =    Hardened light brown coloured scab

G =      Glossy skin

?e =     Adverse reaction prevented accurate evaluation of erythema

?od =   Adverse reaction prevented accurate evaluation of oedema

Table 3              Individual Bodyweights and Bodyweight Changes

Rabbit Number
and Sex

Individual Bodyweight (kg)

Bodyweight Change (kg)

Day 0

Day 14

70905Male

2.28

2.53

0.25

70983Male

2.55

2.76

0.21


Applicant's summary and conclusion

Interpretation of results:
moderately irritating
Remarks:
Migrated information Criteria used for interpretation of results: other: Draize
Conclusions:
The test item produced a primary irritation index of 4.0 and was classified as a MODERATE IRRITANT to rabbit skin according to the Draize classification scheme. No corrosive effects were noted.
The test item was classified as irritant according to EU labelling regulations Commission Directive 2001/59/EC. The symbol “Xi”, the indication of danger “Irritant” and the risk phrase R 38 “IRRITATING TO SKIN” are therefore required.
Due to the persistence of inflammation, the test item was classified Category 2: Irritant according to Regulation (EC) No 1272/2008. The Signal Word “Warning” and Hazard Statement “H315: Causes skin irritation” are therefore required.
Executive summary:

Introduction. 

The study was performed to assess the irritancy potential of the test item to the skin of the New Zealand White rabbit. The method was designed to be compatible with the following:

OECD Guidelines for the Testing of Chemicals No. 404 “Acute Dermal Irritation/Corrosion” (adopted24 April 2002)

Method B4 Acute Toxicity (Skin Irritation) of Commission Regulation (EC) No. 440/2008

Results.

3-minute and 1-hour semi-occluded applications of the test item to the intact skin of one rabbit produced no corrosive effects.

A single 4-hour, semi-occluded application of the test item to the intact skin of two rabbits produced well-defined erythema and slight oedema. Other skin reactions noted were light brown discolouration of the epidermis, loss of skin elasticity and flexibility, crust formation and a hardened light brown coloured scab. Glossy skin was noted at both treated skin sites at the 14-Day observation. No corrosive effects were noted. On occasions, adverse reactions prevented accurate evaluation of oedema and/or erythema.

Conclusion.

The test item produced a primary irritation index of 4.0 and was classified as a moderate irritant to rabbit skin according to the Draize classification scheme.

The test item was classified as irritant according to EU labelling regulations Commission Directive 2001/59/EC. The symbol “Xi”, the indication of danger “Irritant” and the risk phrase R 38 “IRRITATING TO SKIN” are therefore required.

Due to the persistence of inflammation, the test item was classified Category 2: Irritant according to Regulation (EC) No 1272/2008. The Signal Word “Warning” and Hazard Statement “H315: Causes skin irritation” are therefore required.