Registration Dossier

Administrative data

Endpoint:
dermal absorption in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
08 April 2010 to 05 May 2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Cross-reference
Reason / purpose:
reference to same study
Reference
Endpoint:
skin irritation: in vitro / ex vivo
Remarks:
In Vitro Percutaneous Absorption
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
08 April 2010 to 05 May 2010
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Justification for type of information:
This endpoint study record is part of a Weight of Evidence approach comprising a direct observation from RIPT studies, dermal irritation reported during developmental toxicity studies, irritation reported during an LLNA study and the results of an in vitro dermal adsorption study. The data sources are in agreement regarding dermal irritation and are sufficient to fulfil the information requirements as further explained in the provided endpoint summary.
Reason / purpose:
reference to other study
Reason / purpose:
reference to other study
Reason / purpose:
reference to other study
Reason / purpose:
reference to other study
Reason / purpose:
reference to other study
Reason / purpose:
reference to same study
Guideline:
other: OECD Guideline 428 (Skin Absorption: In Vitro Method)
Version / remarks:
Dermal irritation observations reported
Principles of method if other than guideline:
An OECD Guideline 428 (Skin Absorption: In Vitro Method) study was conducted. The dermal irritation observations are reported.
GLP compliance:
yes
Specific details on test material used for the study:
Specific details on test material used for the study
- Radiolabelling: yes
- Source and lot/batch No.of test material: American Radiolabelled Chemicals Inc, batch no. 090109
- Radiochemical purity: 93.6%
- Specific activity: 5 mCi/mmol
- Locations of the label: carbonyl carbon
Test system:
human skin model
Details on animal used as source of test system:
Six samples of full-thickness human skin (4 abdomen and 2 breast) were obtained from female patients aged 19 to 63 years old attending St. John’s Hospital, NHS Lothian, Livingston, UK. On arrival at Charles River, these samples were cleaned of subcutaneous fat and connective tissue using a scalpel blade. The skin samples were washed in cold running water and dried using tissue paper.

The skin samples were then cut into smaller pieces (where appropriate), wrapped in aluminium foil, put into self sealing plastic bags and stored at -20°C until they were used in the study.
Vehicle:
other: leave-on hair styling cream
Amount/concentration applied:
The leave-on hair styling cream containing 2% (w/v) radiolabelled test substance (concentration by radioactivity 2.11% w/v) was applied, at an application rate of about 20 mg/cm² (469 μg equiv. OFPMA/cm²)
Duration of treatment / exposure:
24 hours (single exposure)
Number of replicates:
24 human split thickness skin membranes (12 occluded and 12 unoccluded)
Type of coverage:
other: occluded and unoccluded skin were assessed
Vehicle:
other: leave-on hair styling cream
Details on study design:
SKIN PREPARATION
- Source of skin: female patients aged 19 to 63 years old attending St. John’s Hospital, NHS Lothian, Livingston, UK
- Type of skin: breast or abdomen
- Preparative technique: Split-thickness membranes were prepared by pinning the full-thickness skin, stratum corneum uppermost, onto a raised cork board and cutting at a setting equivalent to 200-400 μm depth using a Zimmer® electric dermatome.
- Thickness of skin (in mm): full-thickness 1430 to 2130 μm; split-thickness 400 μm
- Membrane integrity check: A tritiated water barrier integrity test was performed and any human skin sample exhibiting absorption greater than 0.6% of the applied dose was excluded from subsequent absorption measurements.
- Storage conditions: The split-thickness membranes were stored at about -20°C.

PRINCIPLES OF ASSAY
- Diffusion cell: automated flow-through diffusion cell apparatus
- Receptor fluid: Phosphate buffered saline containing polyoxyethylene 20-oleyl ether (PEG, about 6%, w/v), sodium azide (about 0.01%), streptomycin (about 0.1 mg/mL) and penicillin G (about 100 units/mL)
- Solubility od test substance in receptor fluid: solubility of OFPMA in phosphate buffered saline is 900 mg/L
- Flow-through system: flow rate 1.451 to 1.571 mL/h
- Test temperature: 31.9 to 32.8 °C
- Occlusion: For twelve of the cells (Test Group 1), the donor chambers were occluded with an occlusive trap containing carbon filters. For the other twelve cells (Test Group 2), the donor chamber was left open to the atmosphere and the system was used inside a fume hood.
Remarks on result:
not measured/tested
Other effects / acceptance of results:
Following topical application of [14C]-OFPMA in the test preparation (2%, w/v) to occluded skin, the absorbed dose and dermal delivery of [14C]-OFPMA were 1.18% (5.52 μg equiv./cm2) and 1.53% (7.18 μg equiv./cm2), respectively. There was no report of damage to the stratum corneum.
Interpretation of results:
study cannot be used for classification
Remarks:
Weight of Evidence classification: GHS criteria not met
Conclusions:
Following topical application of [14C]-OFPMA in the test preparation (2%, w/v) to occluded skin, the absorbed dose and dermal delivery of [14C]-OFPMA were 1.18% (5.52 μg equiv./cm2) and 1.53% (7.18 μg equiv./cm2), respectively.  There was no report of damage to the stratum corneum.
Executive summary:

The stratum corneum was observed during a skin irritation observation during a OECD Guideline 428 (Skin Absorption: In Vitro Method) and considered as part of the overall weight of evidence for skin irritation. The in vitro dermal absorption of OFPMA was determined according to OECD Guideline 428 and the accompanying OECD Guidance Document No. 28. Split-thickness human skin membranes were mounted into flow-through diffusion cells. The test preparation containing [14C]-OFPMA (2%, w/v) was applied, at an application rate of about 20 mg/cm² (469 μg equiv. OFPMA/cm²), to 24 human split-thickness skin membranes mounted into flow through diffusion cells in vitro. Immediately after dosing, the donor chamber of 12 cells was covered with an occlusive trap containing carbon filters in an attempt to collect any volatile OFPMA. The remaining 12 cells remained unoccluded and so were open to the atmosphere, simulating the intended consumer use. The integrity of the stratum corneum was observed prior to assessment of the absorbed dose. The Percutaneous absorption study continued to determine dermal adsorption as described elsewhere within this dossier.

 

There was no report of damage to the stratum corneum.

Data source

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

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 428 (Skin Absorption: In Vitro Method)
Version / remarks:
(2004) and the accompanying OECD Guidance Document No. 28 Guidance Document for the Conduct of Skin Absorption Studies (2004)
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Specific details on test material used for the study:
- Source and lot/batch No.of test material: American Radiolabelled Chemicals Inc, batch no. 090109
- Radiochemical purity: 93.6%
- Specific activity: 5 mCi/mmol
- Locations of the label: carbonyl carbon
Radiolabelling:
yes

Test animals

Species:
other: full-thickness human skin
Details on test animals and environmental conditions:
Six samples of full-thickness human skin (4 abdomen and 2 breast) were obtained from female patients aged 19 to 63 years old attending St. John’s Hospital, NHS Lothian, Livingston, UK. On arrival at Charles River, these samples were cleaned of subcutaneous fat and connective tissue using a scalpel blade. The skin samples were washed in cold running water and dried using tissue paper.

The skin samples were then cut into smaller pieces (where appropriate), wrapped in aluminium foil, put into self sealing plastic bags and stored at -20°C until they were used in the study.

Administration / exposure

Type of coverage:
other:
Remarks:
occluded and unoccluded skin were assessed
Vehicle:
other: leave-on hair styling cream
Duration of exposure:
24 hours
Doses:
The leave-on hair styling cream containing 2% (w/v) radiolabelled test substance was applied (concentration by radioactivity was 2.11% w/v), at an application rate of about 20 mg/cm² (469 µg equiv. OFPMA/cm²).
Details on in vitro test system (if applicable):
SKIN PREPARATION
- Source of skin: female patients aged 19 to 63 years old attending St. John’s Hospital, NHS Lothian, Livingston, UK
- Type of skin: breast or abdomen
- Preparative technique: Split-thickness membranes were prepared by pinning the full-thickness skin, stratum corneum uppermost, onto a raised cork board and cutting at a setting equivalent to 200-400 μm depth using a Zimmer® electric dermatome.
- Thickness of skin (in mm): full-thickness 1430 to 2130 µm; split-thickness 400 µm
- Membrane integrity check: A tritiated water barrier integrity test was performed and any human skin sample exhibiting absorption greater than 0.6% of the applied dose was excluded from subsequent absorption measurements.
- Storage conditions: The split-thickness membranes were stored at about -20°C.

PRINCIPLES OF ASSAY
- Diffusion cell: automated flow-through diffusion cell apparatus
- Receptor fluid: Phosphate buffered saline containing polyoxyethylene 20-oleyl ether (PEG, about 6%, w/v), sodium azide (about 0.01%), streptomycin (about 0.1 mg/mL) and penicillin G (about 100 units/mL)
- Solubility of test substance in receptor fluid: solubility of OFPMA in phosphate buffered saline is 900 mg/L
- Flow-through system: flow rate 1.451 to 1.571 mL/h
- Test temperature: 31.9 to 32.8 °C
- Occlusion: For twelve of the cells (Test Group 1), the donor chambers were occluded with an occlusive trap containing carbon filters. For the other twelve cells (Test Group 2), the donor chamber was left open to the atmosphere and the system was used inside a fume hood.

Results and discussion

Total recovery:
The mean mass balance of the occluded test group was 96.01% (standard deviation of 6.25%) of the applied radioactivity.

The mean mass balance of the unoccluded test group was 6.21% (SD = 0.51%) of the applied radioactivity. This very low mass balance was considered to be as a result of the test item volatility.
Percutaneous absorptionopen allclose all
Key result
Time point:
24 h
Dose:
469 µg equiv. OFPMA/cm²
Parameter:
rate
Absorption:
1.53 %
Remarks on result:
other: Dermal Delivery (sum of absorbed dose, epidermis and dermis)
Remarks:
(7.18 μg equiv./cm²; occluded sample)
Time point:
24 h
Dose:
469 µg equiv. OFPMA/cm²
Parameter:
rate
Absorption:
1.18 %
Remarks on result:
other: Absorbed dose (sum of the receptor fluid and receptor rinse)
Remarks:
(5.52 μg equiv./cm²; occluded sample)
Time point:
24 h
Dose:
469 µg equiv. OFPMA/cm²
Parameter:
rate
Absorption:
0.49 %
Remarks on result:
other: Dermal Delivery (sum of absorbed dose, epidermis and dermis)
Remarks:
(2.32 μg equiv./cm²; unoccluded sample)
Time point:
24 h
Dose:
469 µg equiv. OFPMA/cm²
Parameter:
rate
Absorption:
0.18 %
Remarks on result:
other: Absorbed dose (sum of the receptor fluid and receptor rinse)
Remarks:
(0.87 μg equiv./cm²; unoccluded sample)

Any other information on results incl. tables

For the occluded sample, the total dislodgeable dose was 93.25% of the applied radioactivity. The mean total unabsorbed dose was 94.48% of the applied radioactivity. The absorbed dose (1.18%) was the sum of the receptor fluid (1.17%) and the receptor rinse (<0.01%). Dermal delivery (1.53%) was the sum of the absorbed dose, the epidermis (0.19%) and the dermis (0.17%). Steady state absorption was not observed over the 24 hour assessment period and, therefore, a lag time could not be calculated.

 

For the unoccluded sample, the total dislodgeable dose was 5.02% of the applied radioactivity. The mean total unabsorbed dose was 5.71% of the applied radioactivity. The absorbed dose (0.18%) was the sum of the receptor fluid (0.18%) and the receptor rinse (<0.01%). Dermal delivery (0.49%) was the sum of the absorbed dose, the epidermis (0.12%) and the dermis (0.19%). Steady state absorption was not observed over the 24 h assessment period and, therefore, a lag time could not be calculated.

Applicant's summary and conclusion

Conclusions:
Following topical application of [14C]-OFPMA in the test preparation (2%, w/v) to occluded skin, the absorbed dose and dermal delivery of [14C]-OFPMA were 1.18% (5.52 μg equiv./cm2) and 1.53% (7.18 μg equiv./cm2), respectively. The total dislodgeable dose was 93.25%. The mass balance was complete (96.01%). Steady state absorption was not observed over the 24 hour assessment period and, therefore, a lag time could not be calculated.
Executive summary:

The in vitro dermal absorption of OFPMA was determined according to OECD Guideline 428 and the accompanying OECD Guidance Document No. 28. Split-thickness human skin membranes were mounted into flow-through diffusion cells. Receptor fluid was pumped underneath the skin at a flow rate of about 1.5 mL/h. The skin surface temperature was maintained at about 32°C throughout the experiment. A tritiated water barrier integrity test was performed and any human skin sample exhibiting absorption greater than 0.6% of the applied dose was excluded from subsequent absorption measurements.

 

The test preparation containing [14C]-OFPMA (2%, w/v) was applied, at an application rate of about 20 mg/cm² (469 μg equiv.OFPMA/cm²), to 24 human split-thickness skin membranes mounted into flow-through diffusion cells in vitro. Immediately after dosing, the donor chamber of 12 cells was covered with an occlusive trap containing carbon filters in an attempt to collect any volatile OFPMA. The remaining 12 cells remained unoccluded and so were open to the atmosphere, simulating the intended consumer use.

 

Percutaneous absorption was assessed by collecting receptor fluid in hourly fractions from 0 to 8 hours post application and then in 2-hourly fractions from 8 to 24 hours post application. At 24 hours post application, carbon traps were removed from the occluded cells and exposure was terminated by washing the skin surface of all 24 samples. The skin was then dried with tissue paper swabs. The underside of the skin was rinsed with receptor fluid. The skin was then removed from the flow-through diffusion cells, dried and the stratum corneum was removed with 20 successive tape strips. The remaining skin was divided into exposed and unexposed skin (i.e. the area of skin under the cell flange). The exposed epidermis was separated from the dermis. All skin samples were solubilised with Soluene-350® tissue solubiliser. All samples were analysed by liquid scintillation counting.

 

The absorbed dose and dermal delivery of [14C]-OFPMA under occluded conditions were 1.18% (5.52 μg equiv./cm²) and 1.53% (7.18 μg equiv./cm²), respectively. The total dislodgeable dose was 93.25%. The mass balance was complete (96.01%). For unoccluded skin, representing the intended consumer use, the absorbed dose and dermal delivery of [14C]-OFPMA were 0.18% (0.87 μg equiv./cm²) and 0.49% (2.32 μg equiv./cm²), respectively. The total dislodgeable dose was 5.02%. The mass balance was only 6.21% and losses were considered to be due to [14C]-OFPMA volatility. Volatility was confirmed by comparison with the results obtained under occlusive conditions. Steady state absorption was not observed over the 24 hour assessment period for either the occluded or unoccluded systems and, therefore, a lag time could not be calculated.