N-(1,1,3,3-tetramethylbutyl)acrylamide

Dermal absorption

Administrative data

Endpoint:

dermal absorption in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1998-04-22 to 1998-04-30

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study in accordance with OECD draft guidelines (1996) and ECVAM and COLIPA guidelines

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1998

Report date:

1998

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 428 (Skin Absorption: In Vitro Method)

Version / remarks:

The guideline was adopted in 2004, test was conducted in accordance with OECD TG draft from 1996

Deviations:

no

Principles of method if other than guideline:

Test followed COLIPA guidelines (Cosmetic Ingredients: Guidelines for Percutaneous Penetration/Absorption, November 1995)

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

N-(1,1,3,3-tetramethylbutyl)acrylamide

EC Number:

224-169-7

EC Name:

N-(1,1,3,3-tetramethylbutyl)acrylamide

Cas Number:

4223-03-4

Molecular formula:

C11H21NO

IUPAC Name:

N-(1,1,3,3-tetramethylbutyl)acrylamide

Details on test material:

- Name of test material (as cited in study report): N-tert-octylacrylamide
- Physical state: Off white, waxy solid
- Analytical purity: 99.73%
- Impurities (identity and concentrations): not reported
- Purity test date: not reported
- Lot/batch No.: KGS-1152
- Expiration date of the lot/batch: not reported, but substance was used within expiry date
- Storage condition of test material: room temperature

Radiolabelling:

no

Test animals

Species:

other: Human female (post mortem) and male rat

Strain:

Wistar

Sex:

male

Details on test animals or test system and environmental conditions:

Human skin: female, abdominal skin obtained post mortem
Rat skin: male rats of the Wistar-derived strain (Charles River UK, Margale, Kent, UK) aged 28 ± 2 days

Administration / exposure

Type of coverage:

open

Vehicle:

unchanged (no vehicle)

Duration of exposure:

24 hours

Doses:

10 mg/cm2; total weight applied was 25.4 mg

No. of animals per group:

6 tests with human skin; 6 tests with rat skin

Control animals:

no

Details on in vitro test system (if applicable):

SKIN PREPARATION
- Source of skin: human female skin was obtained post mortem
- Ethical approval if human skin: not reported
- Type of skin: human abdominal
- Preparative technique (human skin): extraneous tissue was removed from whole skin samples; skin samples were immersed in water at 60 °C for about 45 seconds and the epidermis teased off the dermis; epidermal membrane was stored frozen on aluminium foil until use
- Preparation technique (rat skin): skin was obtained from dorsal and flank regions; skin was carefully shaved avoiding damage of the skin and shaved skin was excised and subcutaneous fat removed; removed skins were soaked for 20 hours in 1.5 M sodium bromide and then rinsed in distilled water; epidermis was carefully peeled from the dermis and stored frozen on aluminium foil until use
- Thickness of skin (in mm): not reported
- Membrane integrity check: samples of epidermis were mounted in glass diffusion cells with an exposed area of 2.54 cm2; cells were placed in a water bath maintained at 32 ± 1 °C; membrane integrity was determined by measuring the electrical resistance across the membrane; membranes with < 10 kOhm (human) or < 2.5 kOhm (rat) were rejected
- Storage conditions: frozen on aluminium foil
- Justification of species, anatomical site and preparative technique: skin type, application rates and exposure conditions were related to COLIPA guidelines

PRINCIPLES OF ASSAY
- Diffusion cell: glass chamber consisting of donor chamber, receptor chamber and sampling arm (skin membrane and support grid were placed between the donor and receptor chambers)
- Receptor fluid: 50% ethanol in water
- Solubility of test substance in receptor fluid: not reported
- Static system: yes
- Test temperature: 32 ± 1 °C
- Humidity: not reported
- Occlusion: open
- Reference substance(s): di-n-butyl phthalate, dimethyl phthalate, 2-(2-butoxyethoxy) ethanol, n-butanol, benzyl alcohol, 2-phenyl ethanol, ethanol, 2-ethoxyethyl acetate, 2-methoxyethanol, methanol

Results and discussion

Absorption in different matrices:

- Donour chamber: washed with ethanol at the end of experiment and sample was analysed
- Skin wash: the surface of the epidermis ws rinsed with 5 mL volumes of distilled water, rinsings were combined, made up to a known volume with ethanol and analysed
- Skin test site: the epidermis was carefully removed from the receptor chamber and extracted with ethanol and the extracts were analysed
- Receptor fluid, receptor chamber, donor chamber (in vitro test system): at recorded intervals, 0.5 mL samples of the receptor fluid were taken for analysis

Total recovery:

- Total recovery: 90% for human skin and 90.6% for rat skin
- Recovery of applied dose acceptable: yes
- Results adjusted for incomplete recovery of the applied dose: no
- Limit of detection (LOD): not reported
- Quantification of values below LOD or LOQ: no quantification of values below LOQ

Percutaneous absorption

Percutaneous absorption 1

Dose:

10 mg/cm2

Parameter:

percentage

Absorption:

0.1 %

Remarks on result:

other: 6 to 24 hours

Remarks:

Human skin

Percutaneous absorption 2

Dose:

10 mg/cm2

Parameter:

percentage

Absorption:

0.3 %

Remarks on result:

other: 6 to 24 hours

Remarks:

Rat skin

Conversion factor human vs. animal skin:

The ration of percutaneous absorption rates in human and rat skin was 1:2.66

Any other information on results incl. tables

Table 1: Summary of distribution and mass balance of dose

	Percent of dose recovered
Sampled material	Human skin	Rat skin
Donor chamber	4.1	0.6
Skin wash after 24 hours	85.7	89.6
Epidermal membrane	0.1	<0.2
Receptor fluid at 24 hours	0.1	0.3
Total	90.0	90.6

 

Table 2: Absorption of rates determined in single experiments with human and rat epidermis

Diffusion cell number	Skin type and reference	Absorption rate (µg/cm^2/hour)
1	Human, 4217	0.262
2	Human, 4217	0.347
5	Human, 4219	0.342
6	Human, 4219	0.539
9	Human, 4218	1.061
10	Human, 4218	0.581
Mean (n = 6)		0.522
Standard deviation		0.213
12	Rat, 3404	1.924
13	Rat, 3404	1.529
18	Rat, 3406	1.538
19	Rat, 3406	1.108
22	Rat, 3407	0.965
23	Rat, 3407	1.253
Mean (n = 6)		1.386
Standard deviation		0.317

 

Table 3: Absorption rates of comparative organic substances

Substance	Absorption rate (µg/cm^2/h)
Di-n-butyl phthalate	6.6
Dimethyl phthalate	33.0
2-(2-Butoxyethoxy) ethanol	35.0
n-butanol	60.0
Benzyl alcohol	73.0
2-phenyl ethanol	260.0
Ethanol	570.0
2-ethoxyethyl acetate	800.0
2-methoxyethanol	2820.0
Methanol	8400.0*
* Methanol causes extensive damage to the stratum corneum

Applicant's summary and conclusion

Conclusions:

Dermal absorption of N-tert-octylacrylamide through human skin in vitro was found to be very slow when compared with the absorption rates of other organic materials. Further, data from the in vitro test show that the use of rat as model for dermal absorption is likely to over-estimate the absorption of the test substance in human skin.

Executive summary:

The dermal absorption of N-tert-octylacrylamide through human and rat skin was studied under GLP in an in-vitro assay according to COLIPA guidelines (Cosmetic Ingredients: Guidelines for Percutaneous Penetration/Absorption, November 1995), which are equivalent to the principles of OECD TG 428.

For human epidermis, the amounts of subtance absorbed 6, 8 or 10 hours after application were either below or close to the limit of quantification (<5.315, 5.335 and 4.508 µg/cm2). The amount of substance absorbed after 24 hours was 9.397 µg/cm2. The mean absorption rate between 6 and 24 hours after application was 0.522 µg/cm2/hour. The mass balance gave a total mean percentage recovery of 90% of the applied dose. The greatest portion of the recovered dose was readily removed from the skin surface by mild washing (85.7% of applied dose). The epidermal membrane contained only 0.1% of the applied dose.

For rat epidermis, the mean absorption rate of substance over an exposure interval of 24 hours was 1.386 µg/cm2/hour. Steadily increasing amounts of substance were absorbed over periods of 8, 10 an 24 hours with mean amounts reaching 9.371, 12.31 and 31.878 µg/cm2, respectively. The value after 24 hours was equivalent to 0.31% of the applied dose. The mass balance gave a total mean percentage recovery of 90.6% of the applied dose, 90% of which was recovered from the skin surface by mild skin washing. The amounts recovered from the epidermal membrane were below the limit of quantification.

The recovery of applied dose of about 90% for human and rat skin is considered to be acceptable. Dermal absorption of N-tert-octylacrylamide through human and rat skin was demonstrated to be very slow. Rat skin was more permeable than human skin by a factor of 2.66.