In vitro skin penetration and degradation of peptides and their analysis using a kinetic model

The main purpose of this study was to estimate the net percutaneous absorpt ion of physiologically active peptides in vitro. The degradation of two pep tides, Leu-enkephalin (Enk) and Tyr-Pro-Leu-Gly amide (TPLG), during skin p enetration and on the dermal side following penetration, and the prevention of degradation by some protease inhibitors, were investigated using rat sk in in vitro. In addition, these permeation and degradation data were analyz ed using a kinetic model, These peptides were rapidly degraded in the recep tor fluid of a Franz diffusion cell (rate constant: 0.977 h(-1) for Enk and 0.250 h(-1) for TPLG). The addition of phenylmethlsulfonyl fluoride (PMSF) and phenanthroline and the pretreatment of skin with these inhibitors prev ented almost completely and, degradation in the receptor fluid and skin, re spectively. The pretreatment of skin with PMSF and phenanthroline had no ef fect on the penetration of dextran (1000 Da). The degradation rate constant during skin penetration, calculated from the difference in the penetration rate constants via pretreated and untreated skins, was also high (0.037 h( -1) for Enk and 0.050 h(-1) for TPLG). A kinetic model including an input r ate (zero-order), the permeation rate across the viable skin (first-order) and the degradation rate in skin (first-order) was sufficient to describe t he apparent steady-state flux of the peptides through skin. We have, thus, established a method for measuring the true flux of peptides across skin in vitro and a kinetic model which simply describes the skin penetration of p eptides.