Regulatory Toxicology and Pharmacology 35, 198–208 (2002) doi:10.1006/rtph.2001.1530, available online at http://www.idealibrary.com on Comparative in Vitro –in Vivo Percutaneous Penetration of the Fungicide ortho-Phenylphenol Nicole H. P. Cnubben, ∗ Graham R. Elliott,† Betty C. Hakkert,‡ Wim J. A. Meuling,§ and Johannes J. M. van de Sandt ∗ ∗ Department of Explanatory Toxicology, ‡Department of Toxicological Risk Assessment, and §Department of Target Organ Toxicology, TNO Nutrition and Food Research, P.O. Box 360, Utrechtseweg 48, 3700 AJ Zeist, The Netherlands; and †Department of Pharmacology, TNO Prins Maurits Laboratory, P.O. Box 45, 2280 AA Rijswijk, The Netherlands Received May 16, 2001; published online March 28, 2002 The validity of in vitro and in vivo methods for the prediction of percutaneous penetration in humans was assessed using the fungicide ortho-phenylphenol (OPP) (log Po /w 3.28, MW 170.8, solubility in water 0.7 g/L). In vivo studies were performed in rats and human volunteers, applying the test compound to the dorsal skin and the volar aspect of the forearm, respec- tively. In vitro studies were performed using static dif- fusion cells with viable full-thickness skin membranes (rat and human), nonviable epidermal membranes (rat and human), and a perfused pig ear model. For the pur- pose of conducting in vitro/in vivo comparisons, stan- dardized experimental conditions were used with re- spect to dose (120 μg OPP/cm 2 ), vehicle (60% aqueous ethanol), and exposure duration (4 h). In human volun- teers, the potentially absorbed dose (amount applied minus dislogded) was 105 μg/cm 2 , while approximately 27% of the applied dose was excreted with urine within 48 h. In rats these values were 67 μg/cm 2 and 40%, respectively. In vitro methods accurately predicted hu- man in vivo percutaneous absorption of OPP on the ba- sis of the potential absorbed dose. With respect to the other parameters studied (amount systemically avail- able, maximal flux), considerable differences were ob- served between the various in vitro models. In viable full-thickness skin membranes, the amount systemi- cally available and the potentially absorbed dose cor- related reasonably well with the human in vivo sit- uation. In contrast the K p /maximal flux considerably underestimated the human in vivo situation. Although epidermal membranes overestimated human in vivo data, the species differences observed in vivo were re- flected correctly in this model. The data generated in the perfused pig ear model were generally interme- diate between viable skin membranes and epidermal membranes. C  2002 Elsevier Science (USA) Key Words: ortho-phenylphenol; dermal penetration; in vitro–in vivo comparison; risk assessment. INTRODUCTION To predict human internal exposure as a result of dermal exposure, skin absorption data are predomi- nantly obtained from animal studies. In addition, per- cutaneous absorption data obtained from in vitro ex- periments are increasingly used to predict the human in vivo situation (Beck et al., 1994; Ng et al., 1992; Hotchkiss et al., 1992). However, up till now these in vitro models are validated only to a limited extent with data from in vivo experiments. Moreover, in vitro data are often compared to in vivo data from studies ob- tained under different experimental circumstances and exact information on relevant experimental conditions is frequently lacking (ECETOC, 1993). For the in vivo assessment of percutaneous absor- ption, guidelines have been drafted by the OECD (2000a,b) and by the U.S. EPA (1998). Methodology for the in vitro skin absorption studies has been described by various organizations, such as OECD (2000a,b), U.S. EPA (1999), ECETOC (1993), ECVAM (1996), and COL- IPA (Diembeck et al., 1999), but no generally accepted protocol has been developed. In order to study the validity of in vitro and rat in vivo experiments for extrapolation to percutaneous absorp- tion in humans and to characterize possible differences between in vitro and in vivo methods, it is of utmost importance to obtain percutaneous penetration data under identical experimental conditions. Factors like exposure conditions (dose, vehicle, humidity, tempera- ture, and exposure duration) and the type of skin or anatomical site of application (Lotte et al., 1987, 1993; Rougier et al., 1986) are known to influence dermal pen- etration of compounds (ECETOC, 1993). In addition, as dermal penetration also depends on the intrinsic physicochemical characteristics of a compound, differ- ences between in vitro and in vivo methods may depend on parameters such as log Po/w, solubility, and molec- ular weight. 198 0273-2300/02 $35.00 C  2002 Elsevier Science (USA) All rights reserved.