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Skin irritation / corrosion

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Administrative data

Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
02 June 2010 to 07 June 2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report date:
2010

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
other: Method B46 In vitro skin irritation: Reconstituted Human Epidermis Model Test
Deviations:
no
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Chemical structure
Reference substance name:
Magnesium carbonate
EC Number:
208-915-9
EC Name:
Magnesium carbonate
Cas Number:
546-93-0
Molecular formula:
CH2O3.Mg
IUPAC Name:
magnesium carbonate
Details on test material:
- Name of test material (as cited in study report): Magnesium Carbonate
- Physical state: white powder
- Lot No.: 078K0138
- Expiration date of the lot: 30 August 2013
- Storage condition of test material: room temperature in the dark

Test animals

Species:
other: Reconstituted human epidermis model
Strain:
other: Reconstituted human epidermis model
Details on test animals or test system and environmental conditions:
Not applicable

Test system

Type of coverage:
other: Topical
Preparation of test site:
other: Not applicable
Vehicle:
unchanged (no vehicle)
Controls:
other: Dulbecco’s Phosphate Buffered Saline (PBS) with Ca2+ and Mg2+ was used as the negative control. Sodium Dodecyl Sulphate (SDS) was used as the positive control and was prepared as a 5% w/v aqueous dilution.
Amount / concentration applied:
TEST MATERIAL
The test material was used as supplied.
Approximately 10 mg of the test material was applied to the epidermis surface. The epidermis surface had previously been moistened with 5 µL of sterile distilled water to improve contact between the solid test material and the epidermis.
Duration of treatment / exposure:
15 minutes exposure followed by a post exposure incubation period of 42 hours
Observation period:
Not applicable
Number of animals:
Not applicable
Details on study design:
TEST SITE
- Area of exposure: 10 mg of the test material was applied to the epidermis surface.
- % coverage: The test material was applied topically to the corresponding tissues ensuring uniform covering.

REMOVAL OF TEST SUBSTANCE
- Washing: At the end of the exposure period, each tissue was removed from the well using forceps and rinsed using a wash bottle containing PBS with Ca2+ and Mg2+. Rinsing was achieved by filling and emptying each tissue insert for approximately 40 seconds using a constant soft stream of PBS to gently remove any residual test material.
- Time after start of exposure: 15 minutes post exposure

SCORING SYSTEM:
Quantitative MTT Assessment (percentage tissue viability): For the test material the relative mean tissue viabilities obtained after the 15 minute treatment followed by the 42 hour post-exposure incubation period were compared to the mean of the negative control treated tissues (n=3). The relative mean viabilities were calculated in the following way:

(mean OD540 of test material / mean OD540 of negative control) x 100

where OD540 = optical density at 540 nm

Classification of irritation potential is based upon relative tissue viability following the 15 minute exposure period followed by the 42 hour post-exposure incubation period according to the following table:

Criteria for in vitro interpretation Classification
Mean tissue viability is ≤50% Irritant (I) R38
Mean tissue viability is >50% Non-Irritant (NI)

Results and discussion

In vitro

Results
Irritation / corrosion parameter:
other: other: Viability of cells
Value:
98.3
Remarks on result:
other:
Remarks:
Basis: other: Relative mean viability of cells (%). Time point: 15 minutes. Reversibility: other: Not applicable. Remarks: EPISKIN™ reconstituted human epidermis model . (migrated information)

In vivo

Irritant / corrosive response data:
The relative mean viability of the test material treated tissues was 98.3% after the 15 minute exposure period. Following the 15 minute exposure period the test material treated tissues appeared blue which was considered indicative of viable tissue.

The relative mean tissue viability for the positive control treated tissues was ≤40% relative to the negative control treated tissues and the standard deviation value of the percentage viability was ≤20%. The positive control acceptance criterion was therefore satisfied.
The mean OD540 for the negative control treated tissues was ≥0.6 and the SD value of the percentage viability was ≤20%. The negative control acceptance criterion was therefore satisfied.

Any other information on results incl. tables

Table1: Mean OD540Values and Percentage Viabilities for the Negative Control Material, Positive Control Material and Test Material

Material

OD540 of tissues

Mean OD540 of triplicate tissues

±SD of OD540

Relative individual tissue viability (%)

Relative mean viability (%)

± SD of Relative mean viability (%)

Negative Control Material

0.791

0.794

0.024

99.6

100*

3.0

0.772

97.2

0.819

103.1

Positive Control Material

0.052

0.036

0.014

6.5

4.5

1.7

0.029

3.7

0.027

3.4

Test Material

0.817

0.780

0.032

102.9

98.3

4.1

0.757

95.3

0.767

96.6

SD=   Standard deviation

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

Table2: Qualitative Evaluation of Tissue Viability (MTT uptake visual evaluation)

Material

Tissue 1

Tissue 2

Tissue 3

Negative Control Material

-

-

-

Positive Control Material

++

++

++

Test Material

-

-

-

MTT visual scoring scheme
-
          =         blue tissue (viable)
+
         =         blue/white tissue (semi-viable)
++
       =         tissue is completely white (dead)

Applicant's summary and conclusion

Interpretation of results:
not irritating
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The test material, magnesium carbonate was considered to be a non-irritant.