Assessment of percutaneous absorption is of importance to many industrial and scientific fields. The principal application areas are the development of (i) transdermal drug delivery devices to deliver drugs across skin to treat ether local (skin) or systemic diseases; (ii) dermatological formulations in medicine, pharmacy and cosmetics; (iii) safety assessment of cosmetics; and (iv) risk assessment of environmental or occupational hazards. Great efforts have been devoted to develop experimental approaches to measure percutaneous absorption. To date, most of the data on percutaneous absorption have been gained in vitro by diffusion chamber experiments, while in vivo data are commonly obtained from animal experiments via biomonitoring. Bronaugh, R. L. (ed.), Percutaneous absorption: drugs-cosmetics-mechanisms-methodology. 1999, Marcel Dekker, Inc. New York, pp. 123; Schaefer H. Redelmeier T. E. (eds), Skin barrier: principles of percutaneous absorption. 1996, S. Karger AG, Basel, Switzerland, p. 310.
A key function of skin is to provide a barrier that protects the body from foreign substances. Any drug or chemical agent must penetrate the skin's barrier to act ether locally or systemically. Decades of study have established that skin comprises two layers, epidermis and dermis. The epidermis has no capillary blood flow but is made up of several layers of enzymatically active cells, while the dermis in the skin inner layer contains the capillary network that transports the drug or chemical agent to the systemic circulation. The outmost layer of the epidermis is stratum corneum, which comprises layers of keratinized dead cells surrounded by intercellular lipid. Stratum corneum behaves like a passive diffusion barrier. It is responsible for limiting the passage of exogenous chemicals across the skin into the systemic circulation. Agatonovic-Kustrin, S., et al., J. Pharm. Biomed. Anal. 26(2001)241-254.
It is difficult to prepare a large quantity of human skin epidermal membranes for industrial and scientific laboratories. It is conventional practice to use polymeric membranes as skin-imitating barriers to study percutaneous absorption. Skin-imitating membranes differ advantageously from human skin epidermis due to their ready availability, uniformity, tensile strength and chemical purity. Feldstein, M. M., et al., J. Controlled. Release 52 (1998)25-40; Baynes, R. E., et al., Toxicology Industrial Health 16(2000) 225-233. Silastic membranes are the most widely used skin-imitating membranes because of their high permeability comparable with human stratum corneum; and its properties can be modified to simulate skin.
There are mainly two kinds of diffusion chambers currently found in the art, Franz diffusion cell (A) and flow-through diffusion cell (B) as shown in FIG. 1. In these diffusion cells the membrane (a) is placed between two chambers, donor (b) and receptor (c), and the compound in question diffuses from the donor phase through the membrane into the receptor phase. In Franz diffusion cell samples are withdrawn periodically from the receptor phase (g) and analyzed to measure the penetration flux. In the flow-through cell the compounds passing through the membrane are carried away by the receptor fluid flowing beneath the membrane undersurface to be collected in discrete volumes at a remote location. The advantages of the flow-through cell are allowing automatic sampling; maintaining sink conditions since the receptor fluid is replaced continuously, and mimicking the subcutaneous blood flow by the movement of the receptor fluid beneath the undersurface of the membrane. Bronaugh, R. L. (ed.), Percutaneous absorption: drugs-cosmetics-mechanisms-methodology. 1999, Marcel Dekker, Inc. New York, pp. 123; Schaefer H. Redelmeier T. E. (eds), Skin barrier: principles of percutaneous absorption. 1996, S. Karger AG, Basel, Switzerland, p. 310.
The basic compartments of a diffusion cell are illustrated in FIG. 2. The concentration of a given compound is Co in the bulk solution of the donor phase. In the vicinity of the membrane the concentration of the compound is lower than the bulk solution because of the absorption by the membrane, which results in a concentration gradient (Co→Cdx) in the boundary layer between the membrane and the donor phase. There is also a concentration gradient (Crx→Cr) in the boundary layer between the membrane and the receptor phase because the compound passing through the membrane is carried away by the receptor fluid.
The surface concentration in the donor phase (Cdx) is a critical concentration available for the percutaneous absorption. It is determined by the bulk concentration (Co), hydrodynamic agitation, the mass of the compound, and the temperature and viscosity of the solution. At steady-state partition equilibrium is established on both sides of the membrane (Kmd=Cmd/Cdx, Kmr=Cmr/Crx). The surface concentration in the receptor phase (Crx) is also a relevant concentration for the penetration. It is this available concentration that determines the diffusion rate into the receptor phase rather than the concentration in the membrane.
There are several problems in current diffusion cells for in vitro percutaneous absorption. First of all, only trace amounts of compound penetrate the membrane into the receptor phase. A very sensitive Liquid Scintillation Counting (LSC) instrument is required. Thus, chemical agents of interest must often be radiolabeled, and often only one chemical can be studied at a time. This makes the current assessment of percutaneous absorption very expensive and time consuming. Additionally, millions of chemicals in varieties of industrial and environmental matrices are needed to be screened for percutaneous absorption, and many more formulations need to be screened for better drugs, pharmaceuticals, and cosmetics.
Moreover, current techniques using radiolabeled chemical agents cannot handle multiple chemicals and their combinations. Conventional diffusion cells are also designed to mimic an in vivo situation, but experiments are not performed with rigorous compliance to the diffusion laws. The boundary layers in the donor phase and receptor phase are not considered. The diffusion coefficient in the membrane calculated from the absorption flux is only an apparent diffusion coefficient that includes contributions of diffusion resistances from the boundary layers and the membrane. Therefore, the experimental data obtained are only useful to predict the in vivo situation under similar experimental conditions; it cannot be simply interpreted by diffusion laws and cannot be directly analyzed and discussed with kinetic and thermodynamic parameters. For example, the absorption of lipophilic compounds is controlled by the boundary layer diffusion. The absorption rate will be affected by hydrodynamic agitation, solution viscosity and temperature rather than by the structure of the membrane. Misleading interpretation of the experimental data would occur if the existence of the boundary layers is not considered. The present invention addresses these and other problems in the art.