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Toxicological information

Dermal absorption

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Administrative data

Endpoint:
dermal absorption
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Well documented publication

Data source

Reference
Reference Type:
publication
Title:
In vitro and in vivo percutaneous absorption of retinol from cosmetic formulations: Significance of the skin reservoir and prediction of systemic absorption
Author:
Yourick et al
Year:
2008
Bibliographic source:
Toxicol. Appl. Pharmacol. ; 231(1); 117-121; 2008

Materials and methods

Principles of method if other than guideline:
According to:
Bronaugh et al.; 1991; In vitro percutaneous absorption studies: Principles, Fundamentals and Application; Boca Raton; CRC Press.
Kraeling et al.; 1997; In vitro percutanous absorption of alpha hydroxy acids in human skin; J. Soc. Cosmet. Chem.; 48; 187-197
GLP compliance:
not specified

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): 3H-Retinol
- Analytical purity: >99%
- Radiochemical purity: >99%
- Specific activity (if radiolabelling): 47 Ci/mmol


Radiolabelling:
yes

Test animals

Species:
other: human/rat
Strain:
other: Rats: Harlan Sprague Dawley (Hsd:Fuzzy-fz)
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Sprague Dawley, Inc, Indianapolis, IN
- Age at study initiation: 3-14 month
- Housing: Polycarbonate shoe box cages on Sani Chip
- Individual metabolism cages: yes
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: min 3 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21+/-1
- Humidity (%): 50 +/- 10
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12




Administration / exposure

Type of coverage:
other: in vivo study: occlusive
Vehicle:
other: Hydroalcoholic gel / oil in water emulsion
Duration of exposure:
24 hours
Doses:
- Nominal doses: 2 mg/cm2
H-retinol was added to 0.3 % (w/w) retinol leading to 0.7 µCi/cell.
No. of animals per group:
In vitro Human: 2 donors (3 replicates)
In vitro Rat: 3 donors (3-4 replicates)
in vivo Rat: 3 animals
Control animals:
no
Details on study design:
VEHICLE

Hydroalcoholic gel (2% hydroxypropylcellulose dissolved in 90% ethanol with 0.1% butylhydroxytoluene (BHT)
Oil in water emulsion (3% polyglyceryl-3-distearate, 3% cetyl stearyl alcohol, 5% propylene glycol, 10% light
mineral oil, 78% distilled water, 0.5% methyl-p-hydroxybenzoate, 0.5% propyl-phydroxybenzoate, and 0.1% BHT)


REMOVAL OF TEST SUBSTANCE

- Washing procedures and type of cleansing agent:
Washing three times with 0.1 ml of a 10% (v/v) liquid detergent solution that was pipetted onto the skin surface. The skin surface was
gently rubbed with cotton-tipped swabs to remove the detergent solution. The skin was rinsed two times with 0.2 ml of distilled water.
- Time after start of exposure: 24hours


SAMPLE COLLECTION
In vitro:
A fraction collector was used to collect receptor fluid as 6-h fractions for a total of 24 or 72 h.
All cotton swab tips were collected in a scintillation vial.
At the end of the study (24 or 72 h), the skin was removed from the diffusion cell and the amount of retinol remaining in the skin was determined.
In vivo:
All washes, the skin dosing site, remaining carcass, aliquots of the urine, feces were collected.


SAMPLE PREPARATION
In vitro:
Skin discs were tape stripped 10-times to remove the stratum corneum, thus producing a viable epidermal/
dermal disc. Each tape strip was placed into a scintillation vial. Skin discs containing the viable epidermis/dermis were then frozen for skin
content analysis later. Skin discs were thawed and homogenized on ice in HHBSS with a
hand-held homogenizer under cooling. From a total of 4 ml, a 1 ml aliquot was removed and mixed with 3 ml of tissue solubilizer .
The digest was incubated in an oven (approximately 60 °C) overnight or
until the tissue was dissolved. After the skin was dissolved, the amount of radioactivity
was determined by liquid scintillation counting.
In vivo:
Washes were collected to determine the amount of unabsorbed material.
The skin dosing site and remaining carcass were each dissolved in
concentrated (5 M) potassium hydroxide. Aliquots of the urine, feces, and dissolved
carcass were analyzed for radioactivity by liquid scintillation counting. The
skin with the patch still intact was cleaned of fat from the underlying tissue. The patch
was then removed and the remaining glue on the skin was removed with hexanesoaked
cotton swabs. A dermatome was used to create a split-thickness skin section
(180–330 μm). The area containing the dosing site was punched into one or two skin discs.
Each skin disc was tape stripped 10-times to determine the amount of 3H-retinol
remaining in the stratum corneum vs. the epidermis/dermis.

ANALYSIS
- Method type(s) for identification: Liquid scintillation counting
Details on in vitro test system (if applicable):
SKIN PREPARATION
- Source of skin:
Dorsal rat skin was removed and the subcutaneous fat was trimmed away.
Fresh viable human skin was obtained as a result of abdominoplasty procedures
from a local cosmetic surgeon.
- Type of skin: dorsal rat , abdominal human
- Preparative technique:
Human skin was placed on ice in HHBSS and subcutaneous fat was removed.
The skin was then gently cleaned with a 10% soap solution and thoroughly rinsed with distilled water. A split-thickness layer of rat or
human skin was prepared with a dermatome.
- Thickness of skin (in mm): 200–320 μm
- Membrane integrity check:
20-min 3H-water test : Human skin diffusion cells that exhibited a percent of the applied dose of 3H-water absorbed greater than 0.35%
(historical limit) were discarded.


PRINCIPLES OF ASSAY
- Diffusion cell: flow-through diffusion cell system, exposed surface area, 0.64 cm2
- Receptor fluid: HHBSS+4% bovine serum albumin+0.001% BHT, pH 7.4
- Flow-through system: Yes (1.5 ml/h)
- Test temperature: 32 °C
- Other: equilibration for at least 30 min before dosing.

Results and discussion

Percutaneous absorptionopen allclose all
Dose:
2 mg/cm2
Parameter:
percentage
Absorption:
2.4 %
Remarks on result:
other: 24 hours
Remarks:
human in vitro (gel). Stratum corneum not included.
Dose:
2 mg/cm2
Parameter:
percentage
Absorption:
4.3 %
Remarks on result:
other: 24 hours
Remarks:
human in vitro (oil-in-water emulsion). Stratum corneum not included.
Dose:
2 mg/cm2
Parameter:
percentage
Absorption:
24.9 %
Remarks on result:
other: 24 hours
Remarks:
rat in vitro (gel). Stratum corneum not included.
Dose:
2 mg/cm2
Parameter:
percentage
Absorption:
28.7 %
Remarks on result:
other: 24 hours
Remarks:
rat in vitro (oil-in-water emulsion). Stratum corneum not included.
Dose:
2 mg/cm2
Parameter:
percentage
Absorption:
16.3 %
Remarks on result:
other: 24 hours
Remarks:
rat in vivo (gel). Stratum corneum not included.
Dose:
2 mg/cm2
Parameter:
percentage
Absorption:
20.6 %
Remarks on result:
other: 24 hours
Remarks:
rat in vivo (oil-in-water emulsion). Stratum corneum not included.

Any other information on results incl. tables

In vitro percutaneous absorption of retinol in human skin using gel and oil-in-water emulsion vehicles

Recovery site

             Gel (24h)

           Gel (72h)

     Emulsion (24h)

      Emulsion (72h)

 

mean % of applied dose

(+/-SE)

mean % of applied dose

(+/-SE)

mean % of applied dose

(+/-SE)

mean % of applied dose

(+/-SE)

Receptor fluid

0.3

0.1

0.5

0.01

1.3

0.1

2.2

0.2

Stratum corneum

3.5

0.4

2.8

0.8

5.9

1.4

4.8

0.8

Viable skin

2.1

1.2

1.0

0.1

3.0

0.6

2.9

0.6

Recovery

87.3

6.3

95.9

0.2

94.8

2.6

96.3

5.3

In vitro percutaneous absorption of retinol in fuzzy rat skin using gel and oil-in-water emulsion vehicles

Recovery site

              Gel (24h)

            Gel (72h)

      Emulsion (24h)

      Emulsion (72h)

 

mean % of applied dose

(+/-SE)

mean % of applied dose

(+/-SE)

mean % of applied dose

(+/-SE)

mean % of applied dose

(+/-SE)

Receptor fluid

6.0

2.3

12.9

3.9

6.5

1.8

15.7

3.3

Stratum corneum

4.2

1.0

3.3

0.5

3.8

0.5

2.8

0.4

Viable skin

18.9

1.9

14.6

2.4

22.2

1.4

16.2

3.1

Recovery

93.6

5.2

98.5

2.0

91.0

3.2

99.5

5.2

In vivo percutaneous absorption of retinol in fuzzy rat skin using gel and oil-in-water emulsion vehicles

Recovery site

            Gel (24h)

          Gel (72h)

     Emulsion (24h)

     Emulsion (72h)

 

mean % of applied dose

(+/-SE)

mean % of applied dose

(+/-SE)

mean % of applied dose

(+/-SE)

mean % of applied dose

(+/-SE)

Total systemic absorption (urine, faeces, carcass)

4.1

0.4

4.4

0.03

6.5

0.2

5.6

0.1

Stratum corneum

5.5

1.3

2.6

0.7

3.4

0.3

7.5

1.4

Total in skin

17.7

2.1

12.9

2.7

17.5

0.7

11.0

1.7

Recovery

83.1

4.5

70.7

2.3

77.9

1.5

81.9

1.0

A higher recovery was found, i.e 106.2% (Gel, 24h), when charcoal filter paper was placed on top of the protective patch around the edges of the dosing area for 1h after test substance application, indicating evaporation of retinol from the dosing site.

Applicant's summary and conclusion