In vitro predictions of skin absorption of caffeine, testosterone, and benzoic acid: a multi-centre comparison study.

To obtain better insight into the robustness of in vitro percutaneous absorption methodology, the intra- and inter-laboratory variation in this type of study was investigated in 10 European laboratories. To this purpose, the in vitro absorption of three compounds through human skin (9 laboratories) and rat skin (1 laboratory) was determined. The test materials were benzoic acid, caffeine, and testosterone, representing a range of different physico-chemical properties. All laboratories performed their studies according to a detailed protocol in which all experimental details were described and each laboratory performed at least three independent experiments for each test chemical. All laboratories assigned the absorption of benzoic acid through human skin, the highest ranking of the three compounds (overall mean flux of 16.54+/-11.87 microg/cm(2)/h). The absorption of caffeine and testosterone through human skin was similar, having overall mean maximum absorption rates of 2.24+/-1.43 microg/cm(2)/h and 1.63+/-1.94 microg/cm(2)/h, respectively. In 7 out of 9 laboratories, the maximum absorption rates of caffeine were ranked higher than testosterone. No differences were observed between the mean absorption through human skin and the one rat study for benzoic acid and testosterone. For caffeine the maximum absorption rate and the total penetration through rat skin were clearly higher than the mean value for human skin. When evaluating all data, it appeared that no consistent relation existed between the diffusion cell type and the absorption of the test compounds. Skin thickness only slightly influenced the absorption of benzoic acid and caffeine. In contrast, the maximum absorption rate of testosterone was clearly higher in the laboratories using thin, dermatomed skin membranes. Testosterone is the most lipophilic compound and showed also a higher presence in the skin membrane after 24 h than the two other compounds. The results of this study indicate that the in vitro methodology for assessing skin absorption is relatively robust. A major effort was made to standardize the study performance, but, unlike in a formal validation study, not all variables were controlled. The variation observed may be largely attributed to human variability in dermal absorption and the skin source. For the most lipophilic compound, testosterone, skin thickness proved to be a critical variable.