Ex vivo percutaneous absorption of solid lipophilic toxic substances: use of an artificial sebum type vehicle

Outline of reasons and objectives
Measuring ex vivo percutaneous absorption of solid lipophilic toxic substances at ambient temperature, such as certain toxic substances already studied before (benzo[a]pyrene, pyrene, naphthalene, bisphenol A, 2,4-dichlorophenoxyacetic acid), is usually performed in solution in a volatile solvent such as acetone. This method, remote from the real conditions of exposure of the skin, frequently suffers from problems of instability of the percutaneous fluxes achieved with the ex vivo skin of human origin or of rat origin. The objective of the study was to find a formulation for artificial sebum whose composition is close to the physiological proportions in humans, and then to compare the ex vivo percutaneous absorption fluxes of the solid substances obtained with the sebum or acetone as a solubilisation vehicle. This study aimed to make it possible to define optimum experimental conditions for better assessing the risk related to exposure to solid lipophilic chemicals via the cutaneous route.
Methodology
In a first stage, bibliographic research made it possible to determine a standard formulation for artificial sebum that was tested by compared measurements of percutaneous absorption flux of a substance that had already been studied. The biocompatibility for the ex vivo skin of the selected formulation was then assessed by using validated methods. Comparison between acetone and the artificial sebum was then performed in humans and in rats by measuring the ex vivo percutaneous absorption fluxes of ten solid lipophilic substances at ambient temperature, with as wide a range of differential solubility (log(kow)) as possible. The variation in ex vivo percutaneous fluxes as a function of skin thickness obtained in rats with sebum were extrapolated for a skin thickness of zero in order to estimate an in vivo percutaneous absorption flux. The results were compared with fluxes obtained with in vivo experimental data from the laboratory achieved with acetone as the vehicle.
Main results
The artificial sebum, made up of squalene (15%), Jojoba oil (25%), triolein (30%), and oleic acid (30%), was stable and non-toxic for the ex vivo skin, with good preservation of the activity of the enzymes of the viable skin. It allowed good solubilisation of all of the tested solid lipophilic substances. The profile of the percutaneous absorption fluxes of the substances as measured for 24 to 48 hours was always constant, without the problems of variation in fluxes encountered with acetone. The percutaneous flux measured with the sebum was greater than or equal to the flux measured with acetone for seven of the ten substances in humans. The ex vivo fluxes obtained with the sebum and extrapolated to a zero skin thickness were all significantly different from the in vivo percutaneous absorption fluxes obtained in rats with the acetone as the vehicle.
Discussion
The main advantages of the artificial sebum compared with the solvents usually used as vehicles for solid lipophilic substances are:
a stable formulation composed of a mixture of four substances that are readily commercially available, and having a substitute close to human sebum;
an absence of toxicity and of irritability of the skin for the artificial sebum, without significantly modifying the activity of the enzymes of the skin in humans and in rats;
unlike with acetone, obtaining a flux profile at equilibrium makes it possible to estimate the equilibrium time of the fluxes and to have a mean flux that is constant for several hours in humans and in rats for the ten tested substances.
The risk of dilipidation of the skin by acetone precludes comparison between the in vivo percutaneous flux measured in rats with acetone and the ex vivo percutaneous flux at zero skin thickness measured with the sebum. This study shows that artificial sebum is an advantageous alternative to using organic volatile solvents as solubilisation vehicles for estimating the risk of exposure of the skin to solid lipophilic toxic substances at ambient temperature.