31P solid-state NMR based monitoring of permeation of cell penetrating peptides into skin

The main objective of the current study was to investigate penetration of cell penetrating peptides (CPPs: TAT, R-8, R-11, and YKA) through skin intercellular lipids using P-31 magic angle spinning (MAS) solid-state NMR. In vitro skin permeation studies were performed on rat skin, and sections (0-60, 61-120, and 121 180 mu m) were collected and analyzed for P-31 NMR signal. The concentration-dependent shift of 0, 25, 50, 100, and 200 mg/ml of TAT on skin layers, diffusion of TAT, R-8, R-11, and YKA in the skin and time dependent permeation of R-11 was measured on various skin sections using P-31 solid-state NMR. Further, CPPs and CPP-tagged fluorescent dye encapsulate liposomes (FLip) in skin layers were tagged using confocal microscopy. The change in P-31 NMR chemical shift was found to depend monotonically on the amount of CPP applied on skin, with saturation behavior above 100 mg/ml CPP concentration. R-11 and TAT caused more shift in solid-state NMR peaks compared to other peptides. Furthermore, NMR spectra showed R-11 penetration up to 180 mu m within 30 min. The results of the solid-state NMR study were in agreement with confocal microscopy studies. Thus, P-31 solid-state NMR can be used to track CPP penetration into different skin layers. (C) 2013 Elsevier B.V. All rights reserved.