Imidazole

Dermal absorption

Administrative data

Endpoint:

dermal absorption in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliant, guideline study, available as unpublished report, no restrictions, fully adequate for assessment

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 428 (Skin Absorption: In Vitro Method)

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: OECD Guidance Document No. 28 for the conduct of skin absorption studies, March 2004

GLP compliance:

yes (incl. QA statement)

Remarks:

BASF SE Experimental Toxicology and Ecology, 67056 Ludwigshafen, Germany

Test material

Test material information

Constituent 1

Reference substance name:

Imidazole

EC Number:

206-019-2

EC Name:

Imidazole

Cas Number:

288-32-4

Molecular formula:

C3H4N2

IUPAC Name:

1H-imidazole

Details on test material:

- Name of test material: 14C-Imidazol
- Storage condition of test material: freezer

- Name of test material: Imidazol
- Storage condition of test material: ambient(RT)
- Physical state: white, solid

Radiolabelling:

yes

Remarks:

2-C14

Test animals

Species:

other: in vitro

Administration / exposure

Vehicle:

water

Duration of exposure:

8 hours

Doses:

- Nominal doses: 100 and 1000 µg/cm²
- Actual doses: (mean ± SD) 99 ± 3 and 930 ± 18 µg/cm²

Details on study design:

NUMBER OF DIFFUSION CELLS:
- 8 cells for each dose

DOSE PREPARATION
- A respective aliquot of the radiolabeled test substance was taken and the organic solvent (ethanol) was evaporated. An adequate amount of the non-labeled test substance and tap water was added to the dried residue. The preparations were stirred in order to guarantee homogeneity.

APPLICATION OF DOSE:
- Single topical application. The test-substance preparation was distributed uniformly on the exposed skin preparation using a displacement pipette.
- Amount(s) applied: about 10 μL / cm²
- Actual doses calculated as follows: The exact amount applied per diffusion cell was determined by reweighing the pipette after application, the weight of which had been previously determined after filling.
- Concentration: 10 and 100 mg/mL
- Exposure period: 8 hours

TEST SITE
- Area of exposure: 1 cm²

SAMPLE COLLECTION FOR PENETRATION RATE
- Aliquots of the receptor medium were automatically sampled by the fraction collector 1, 2, 4, 6, 8, 12, and 24 hours after application

SAMPLE COLLECTION FOR MASS BALANCE
- After several washings and the skin surface had dried, the stratum corneum was removed by tape stripping. Scotch Crystal Clear Tape 600 (Art. No: 11265, 3M, France) Six tape strips were taken. The tapes were pooled into two samples for analysis: the first sample containing the first and second tape and the second samplecontaining the third until sixth tape. The remaining skin was analyzed separately.

TEST SAMPLE STORAGE
- Until analysis the samples were stored at room temperature and extracted, likewise at room temperature, using mechanical agitation and/or ultrasonic treatments.

TEST SAMPLE ANALYSIS
- Weighed aliquots of each native receptor fluid sample or of the solvents used for extraction of the diffusion cell parts were analyzed by liquid scintillation counting (Wallac type 1409 or Perkin Elmer Tri-Carb 2800TR). After addition of scintillation cocktail (Hionic Fluor, Perkin Elmer) the samples were counted for 10 min in a LSC and the disintegration rate was determined.
- The recovery of the test substance applied in the various compartments was determined as non-absorbed fraction (donor chamber, dislodgeable test substance, charcoal filter, fraction present in the tape strips), fraction in the remaining skin preparation and absorbed fraction (contents of the receptor chamber, receptor chamber washing and samples taken).

Details on in vitro test system (if applicable):

SKIN PREPARATION
- Source of skin: Surgically removed skin from abdomen (BIOPREDIC International, Saint-Grégoire, France and Across Barriers GmbH, Science Park 1, Saabrücken, Germany)
- Type of skin: Dermatomed human skin preparations
- Preparative technique: skin preparation was hydrated in physiological saline for about 10 minutes before mounting to the diffusion cells
- Thickness of skin (in mm): 0.299 – 0.419
- Membrane integrity check: Only visually intact skin with a TEER (impedance value) above 1 kOhm and a TEWL below 15 g/m2h were used.
- Storage conditions: Maximum storage time of 3 days in a refrigerator or 12 months at - 20°C.

PRINCIPLES OF ASSAY
- Diffusion cell: Modified Franz cell (Laboratory Glass Apparatus Inc, U.S.A. or BASF SE) consisting of an upper donor chamber and a wide opening at the top and a lower receptor chamber
- Receptor fluid: tap water for the high and low dose
- Solubility of test substance in receptor fluid: The maximum solubility of the test substance in water is 633 g/L.
- Static system: yes
- Test temperature: 32 ± 1°C
- Occlusion: The openings of the donor chambers were covered with a charcoal filter and Fixomull® Stretch adhesive fleece (semi-occlusive conditions)

Results and discussion

Signs and symptoms of toxicity:

not examined

Dermal irritation:

not examined

Absorption in different matrices:

Dose group: 930 μg/cm² (mean amount of test substance [μg] and (mean % of applied dose))
- Total non-absorbed dose: 56.19 µg (6.04%)
- Absorbed: Sum receptor samples 0 - 24 h including wash out: 568.05 μg (61.06%)
- Absorbed: Receptor fluid: 264.41 μg (28.43%)
- Absorbed: Receptor chamber washing: 24.09 μg (2.59%)
- Absorbed: Total absorbed: 856.55 μg (92.09%)
- Total dose associated to skin: 10.65 µg (1.15%)
- The mean absorption rate of all cells corresponding to the steepest slope of the cumulative absorbed dose curve over time was 255.3 μg/(cm²×h). Taking the applied concentration into account, this is equivalent to a permeability constant of 258.6E-5 cm/h. Furthermore the evaluation of the cumulative absorption curves indicated a mean lag time of about 1.07 hours. The cumulative absorbed dose found in the receptor fluid after the 24-hour exposure period was 92.6% of the applied dose.

Dose group: 99 μg/cm² (mean amount of test substance [μg] and (mean % of applied dose))
- Total non-absorbed dose: 23.38 µg (23.59%)
- Absorbed: Sum receptor samples 0 - 24 h including wash out: 38.54 μg (38.94%)
- Absorbed: Receptor fluid: 26.38 μg (26.61%)
- Absorbed: Receptor chamber washing: 2.09 μg (2.11 %)
- Absorbed: Total absorbed: 67.00 μg (67.66 %)
- Total dose associated to skin: 1.98 µg (2.00%)
- The mean absorption rate of all cells corresponding to the steepest slope of the cumulative absorbed dose curve over time was 14.0 μg/cm²×h. Taking into account the applied concentration, this is equivalent to a permeability constant of 142.4E-5 cm/h. Furthermore the evaluation of the cumulative absorption curves indicated a mean lag time of about 1.37 hours. The cumulative absorbed dose found in the receptor fluid after the 24-hour exposure period was 66.2% of the applied dose.

Total recovery:

Dose group: 930 μg/cm²
- The mean total recovery measured in diffusion cells equipped with human skin at the high dose fulfilled the OECD quality criteria. The individual values ranged between 96.17 and 100.62 %

Dose group: 99 μg/cm²
- The mean total recovery measured in diffusion cells equipped with human skin at the low dose fulfilled the OECD quality criteria. The individual values ranged between 90.19 and 97.22 %.

Percutaneous absorption

Percutaneous absorption 1

Dose:

930 μg/cm²

Parameter:

percentage

Absorption:

92.09 %

Remarks on result:

other: 24 hours

Remarks:

8 hour exposure

Percutaneous absorption 2

Dose:

99 μg/cm²

Parameter:

percentage

Absorption:

67.66 %

Remarks on result:

other: 24 hours

Remarks:

8 hour exposure

Applicant's summary and conclusion

Conclusions:

Under the test conditions OECD 428 and GLP, the mean absorbed doses were 92.09 and 67.66 % of dose for skin treated with about 1000 μg/cm² and 100 μg/cm², respectively. Mean absorption lag times were 1.07 hours and 1.37 hours for the high and low dose, respectively and demonstrate the presence of a functional barrier in the skin samples used.

Executive summary:

According to OECD guideline 428 and GLP, the diffusion of 14C-Imidazol into and through human skin was assessed by single topical application. Target doses of 1000μg/cm² and 100μg/cm² were applied to split thickness skin preparations mounted on Franz-type diffusion cells. After the exposure time of 8 h and after the sampling period (24 hours after exposure), skin membranes were washed with a mild soap solution and tap water. At the end of the sampling period the test substance was recovered from all compartments of each diffusion cell. The mean total recoveries fulfill the quality criteria put forward in the test guidelines. The mean absorbed doses were 92.09 and 67.66% of dose for skin treated with the high dose and the low dose, respectively. For all dose groups, minor amounts of the test substance were associated with the skin after the exposure period. These amounts accounted to 1.15 and 2.00 % of dose for the high dose and the low dose, respectively. Mean absorption lag times were 1.07 hours and 1.37 hours for the high and low dose, respectively, and demonstrate the presence of a functional barrier in the skin samples used