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Toxicological information

Dermal absorption

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Administrative data

Endpoint:
dermal absorption in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Peer reviewed database, comparable to guideline study with acceptable restrictions
Cross-reference
Reason / purpose:
reference to same study

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
1993

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 428 (Skin Absorption: In Vitro Method)
Principles of method if other than guideline:
Investigation of percutaneous absorption of 4.4'-methylenedianiline (MDA) after topical application to full-thickness skin from male Fischer 344 rats in flow-through diffusion cells, on four separate occasions.
GLP compliance:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Radiolabeled MDA:
- Radiochemical purity: >95%
- Specific activity: act. 56 mCi/mmol
- Locations of the label: [ring14C]4,4 '-methylenedianiline (Amersham)
Radiolabelling:
yes
Remarks:
[ring14C]4,4´-methylenedianiline

Test animals

Species:
rat
Strain:
Fischer 344
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan-OLAC (Oxford, Oxon, UK)
- Diet: CRM pellets, Biosure (Manea, Cambs., UK); ad libitum
- Water: ad libitum

Administration / exposure

Type of coverage:
other: occlusive and open
Vehicle:
ethanol
Duration of exposure:
72 h
Doses:
- Actual doses: 39.4, 20.1, 21.6, 23.6 µg/cm2
- Dose volume: 5-10 µl
No. of animals per group:
4
Details on in vitro test system (if applicable):
SKIN PREPARATION
- Source of skin: dorsal region of male Fischer 344 rats
- Type of skin: full thickness
- Preparative technique: following sacrificion, the dorsal region was shaved with animal clippers and the shaved area was cut out with dissecting scissors. The rat skin sections were placed, epidermal side uppermost, on a plastic dissecting board and circles, 1.7 cm in diameter, were cut out using a circular sharpened steel cutter. Excess subcutaneous tissue was removed with a scalpel.

PRINCIPLES OF ASSAY
- Flow-through diffusion cell system: consisted of seven teflon diffusion cells and a fraction collector (Crown Glass Co Inc., Somerville, NJ, USA), a thermostatically controlled Churchill water circulator and a model 202V/AA16 cassette peristaltic pump (Watson-Marlow, Falmouth, Cornwall, UK). The receptor volume of each diffusion cell was 130 ul.
- Receptor fluid: HEPES buffered Hanks' balanced salt solution supplemented with 0.5% (v/v) gentamicin, continuously flowing
- Test temperature: 32°C
- the skin was placed in the diffusion cells in the flow-through apparatus, and allowed to equilibrate for about 30min.
- aliquots of 5-10 µl [14C]-MDA (17.7-40.6µg/cm2) in ethanol were applied to the exposed skin surface.
- occlusion with teflon caps or left open.
- receptor fluid (1.5 ml) was collected hourly for up to 72h.
- at the end of the experiment (72 hr) the epidermal surface of the skin was washed, and the washings and skin were counted to determine radioactivity remaining on the skin surface.

ANALYSIS
- Liquid scintillation counting

STATISTICAL EVALUATION:
- Student`s t-Test for unpaired data, level of significance p<0.05.

Results and discussion

Absorption in different matrices:
See table 1
Total recovery:
- Total recovery: 94.5 +/-6 %
Percutaneous absorptionopen allclose all
Dose:
20.1-39.4 ug/cm2
Parameter:
percentage
Absorption:
ca. 6.1 %
Remarks on result:
other: 72 h
Remarks:
open
Dose:
20.1-39.4 ug/cm2
Parameter:
percentage
Absorption:
ca. 13.3 %
Remarks on result:
other: 72h
Remarks:
occlusive

Any other information on results incl. tables

Table 1: Recovery of radioactivity at 72h after topical application of [14C] MDA to rat skin in vitro (% of applied dose).

Rat (unoccluded) (n=12) Rat (occluded) (n=16)
Receptor fluid 6.1 ± 2 13.3 ± 4.0*
Skin surface 25.5 ± 6.8 21.5 ± 7.2
Diffusion cell 5.4 ± 4.4 3.9 ± 2.6
Teflon cap - 0.3 ± 0.2
Skin surface 57 ± 5.6 53.1 ± 8.6
Total 94.5 ± 6.0 92.1 ± 6.5

* significantly different from unoccluded value (p<0.05)

A typical absorption profile showed a lag phase of about 2 hr before a rapid increase in absorption, reaching an equilibrium at 72hr (data not shown).

Applicant's summary and conclusion