Registration Dossier

Administrative data

Endpoint:
dermal absorption in vitro / ex vivo
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Study period:
From August 18, 2005 to August 17, 2006
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
Refer to the Quaternary ammonium salts (QAS) category or section 13 for details on the category justification. The study with the read across substance is considered sufficient to fulfil the information requirements as further explained in the provided endpoint summary.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2006
Report Date:
2006

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 428 (Skin Absorption: In Vitro Method)
Deviations:
no
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Specific details on test material used for the study:
- Name of test material: Alkyl(C12-C16)dimethylbenzylammonium chloride (non-radiolabelled ADBAC) and [14C]-ADBAC (radiolabelled ADBAC)
- Analytical purity: non-radiolabelled: 80.5% pure; radiolabelled: 99.7%
- Composition of test material, percentage of components:
- Lot/batch No.: radiolabelled: 990930; non-radiolabelled: J2220459
- Radiochemical purity (if radiolabelling): 99.7%
- Specific activity (if radiolabelling): 25 mCi/mmol; total radioactivity: 1 mCi/mL in etanol
- Storage condition of test material: non-radiolabelled: ambient temperature in dark
Radiolabelling:
yes

Test animals

Species:
human
Details on test animals and environmental conditions:
Human skin membranes, in vitro

Administration / exposure

Type of coverage:
other: Automated flow-through diffusion cell system
Vehicle:
water
Doses:
0.03% (w/w) and 0.3% (w/w), in water
Details on in vitro test system (if applicable):
Skin preparation:
- Source of skin: Eight samples from Plastic Surgery unit, St. Johns Hospital, West Lothian NHS Trust, Livingston, UK
- Ethical approval if human skin: Yes
- Type of skin: Full-thickness human skin (1 upper arm, 2 abdomen and 5 breast)
- Preparative technique: Skin was transferred to Charles River Laboratories on ice and cleaned of subcutaneous fat and connective tissue using scalpel. Skin was washed in cold running tap water and dried using tissue paper. Each sample was then cut into smaller pieces, wrapped in aluminium foil, put into self sealing plastic bags and stored at -20°C until required. 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.
Principles of assay:
- Diffusion cell: Automated flow-through diffusion cell system (Scott/Dick, University of Newcastle-upon-Tyne, UK)
- Receptor fluid: Tissue culture medium containing approximately 5% (w/v) bovine serum albumin, 1% (w/v) streptomycin (approximately 0.1 mg/mL), and penicillin G (approximately 100 units/mL)
- Solubility of test substance in receptor fluid:
- Flow-through system: Automated
- Test temperature: The mean temperature 19°C (SD=0.8°C)
- Humidity: The mean relative humidity 35% (SD=0.6%)

Results and discussion

Total recovery:
Refer to Table-1
Percutaneous absorptionopen allclose all
Key result
Dose:
0.03%
Parameter:
percentage
Absorption:
0.05 %
Remarks on result:
other: 24 h
Remarks:
96.80% was not absorbed
Key result
Dose:
0.3%
Parameter:
percentage
Absorption:
0.03 %
Remarks on result:
other: 24 h
Remarks:
94.68% was not absorbed

Any other information on results incl. tables

- Low dose (0.030%):

The mean mass balance was 99.03% of the applied dose (3.09 μg equiv./cm2). The mean dislodgeable dose was 60.53% of the applied dose (1.89 μg equiv./cm2). The mean total unabsorbed dose was 96.80% of the applied dose (3.02 μg equiv./cm2). This consisted of the dislodgeable dose, unexposed skin (0.02%) and the radioactivity associated with the stratum corneum (36.25%). The stratum corneum acted as a good barrier to the test substance as the bulk of the radioactivity (30.26%) was recovered in the outermost 5 tape strips (tape strips 1-5). Considerably less radioactivity was recovered with each of the subsequent 3 groups of tape strips (3.20%, 1.82% and 0.97% in tape strips 6-10, 11-15 and 16-20 respectively), suggesting that the test substance would be sloughed off with the skin in the future. The absorbed dose (0.05%, or <0.01 μg equiv./cm2) was the sum of the receptor fluid (0.05%) and the receptor rinse (<0.01%). Dermal delivery (2.22%, or 0.07 μg equiv./cm2) was the sum of the absorbed dose and the exposed skin (2.18%). There was no apparent lag time and the fluxes ranges from 0.03 to 0.12 ng equiv./cm2/h over the 1 to 24-h exposure period.

 

- High dose (0.300%):

The mean mass balance was 96.84% of the applied dose (29.91 μg equiv./cm2). The dislodgeable dose was 77.87% of the applied dose (24.05 μg equiv./cm2). The mean total unabsorbed dose was 94.68% of the applied dose (29.24 μg equiv./cm2). This consisted of the dislodgeable dose, unexposed skin (0.17%) and the radioactivity associated with the stratum corneum (16.64%). The stratum corneum acted as a good barrier to the test item as the bulk of the radioactivity (10.86%) was recovered in the outermost 5 tape strips (tape strips 1-5). Considerably less radioactivity was recovered with each of the subsequent 3 groups of tape strips (3.11%, 1.71% and 0.96% in tape strips 6-10, 11-15 and 16-20, respectively), again suggesting that the test item would be sloughed off with the skin in the future. The absorbed dose (0.03%, or 0.01 μg equiv./cm2) was the sum of the receptor fluid (0.03%) and the receptor rinse (<0.01%). Dermal delivery (2.16%, or 0.67 μg equiv./cm2) was the sum of the absorbed dose and the exposed skin (2.13%). There was no apparent lag time. There was no apparent lag time and the fluxes ranges from 0.22 to 0.74 ng equiv./cm2/h over the 1 to 24-h exposure period.

Table 1. Summary of recoveries after 24h

Test Preparation

Low Concentration

High Concentration

Target test substance concentration (%, w/w)

0.03

0.30

Test substance concentration by Radioactivity (%, w/w)

0.031

0.306

Test preparation application rate (mg/cm2)a

10.01

10.09

Test substance application Rate (μg equiv./cm2)

3.12

30.87

Dislodgeable Dose (% Applied Dose)

60.53

77.87

Unabsorbed Dose (% Applied Dose)

96.80

94.68

Absorbed Dose (% Applied Dose)

0.05

0.03

Dermal Delivery (% Applied Dose)

2.22

2.16

Mass Balance (% Applied Dose)

99.03

96.84

Dislodgeable Dose (μg equiv./cm2)

1.89

24.05

Unabsorbed Dose (μg equiv./cm2)

3.02

29.24

Absorbed Dose (μg equiv./cm2)

<0.01

0.01

Dermal Delivery (μg equiv./cm2)

0.07

0.67

Mass Balance (μg equiv./cm2)

3.09

29.91

a mg of test preparation per cm of skin

 

Applicant's summary and conclusion

Conclusions:
Based on the results of the read across study, following topical application of 14C-radiolabelled test substance in low (0.03%, w/w) and high (0.3%, w/w) concentration, the mean absorbed dose and mean dermal delivery of 14C-radiolabelled test substance were 0.05% (<0.01 μg equiv./cm2) and 2.22% (0.07 μg equiv./cm2) of the applied dose for the low concentration test preparation, respectively, and 0.03% (0.01 μg equiv./cm2) and 2.16% (0.67 μg equiv./cm2) of the applied dose for the high concentration test preparation, respectively. The maximum fluxes for the low and high doses were 0.12 ng equiv./cm2/h and 0.74 ng equiv./cm2/h, respectively, at 2 h.
Executive summary:

A study was conducted to determine the dermal absorption of the read across substance, quaternary ammonium compounds, benzyl C12-C16 (even numbered)-alkyldimethyl chlorides (C12-16 ADBAC),according to OECD Guideline 428, in compliance with GLP. In anin vitrostudy, split-thickness human skin membranes were mounted into flow-through diffusion cells. Receptor fluid was pumped underneath the skin at a flow rate of 1.5 mL/hour. The skin surface temperature was maintained at approximately 32°C. A barrier integrity test using tritiated water was performed and any skin sample exhibiting a permeability coefficient (kp) greater than 2.5 x 10-3 cm/h was excluded from subsequent absorption measurements. Two test preparations containing 14C- radiolabelled test substance (i.e. 0.03% and 0.3%), were applied at an application rate of 10 mg/cm2. Absorption was assessed by collecting receptor fluid in hourly intervals from 0-6 hours post dose and then in 2-hourly intervals from 6-24 h post dose. At 24 h post dose, the exposure was terminated by washing and drying the skin. The stratum corneum was then removed from the skin by 20 successive tape strips. All samples were analysed by liquid scintillation counting. Based on the results of the read across study, following topical application of 14C- radiolabelled test substance in low (0.03%, w/w) and high (0.3%, w/w) concentration test preparations to human skin in vitro, the mean absorbed dose and mean dermal deliveries were 0.05% (0.01 ηg equiv. /cm2) and 2.22% (0.07 ηg equivalent/cm2) of the applied dose for the low concentration test preparation, respectively, and 0.03% (0.01 ηg equivalent /cm2) and 2.16% (0.67 ηg equivalent/cm2) of the applied dose for the high concentration test preparation, respectively. The stratum corneum acted as a barrier to absorption, with the mean total unabsorbed doses (recovered in skin wash, tissue swabs, pipette tips, cell wash, stratum corneum and unexposed skin) of 96.80 and 94.68% of the applied dose for the low and high concentration test preparations, respectively. The maximum fluxes for the low and high doses were 0.12 ηg equivalent /cm2/h and 0.74 ηg equivalent /cm2/h, respectively, at 2 h (Roper, 2006).