*2.6. Tape-Stripping Studies. Drug Penetration through the Skin and Stratum Corneum Depth*

The assay was performed using porcine skin samples. Skin was removed from the connective tissues and placed onto a glass slide, with the outside layer facing upwards. An aluminium mask was placed over it, leaving the application area uncovered. One hundred microliters of each formulation containing lipid vesicles was applied to the delimited application area of skin. The system was then incubated at 32 ◦C for 2, 4, and 6 h. Each test was performed in triplicate (*n* = 3). After incubation, 20 strips of adhesive tape were applied sequentially to the skin using a roller, according to the standardized procedure in our lab, and then removed with a forceps [65]. The amount of stratum corneum removed was determined by infrared densitometry (SquameScan™ 850A device, Wetzlar, Germany), and strips were grouped in different pools, following the sequence: 1, 2, 3–5, 6–10, 11–15, 16–20 [66]. The B12 amount present in each sample was extracted overnight from the strips using a methanol:water mixture (50:50 *v/v*) as extraction solvent, since it afforded an optimal B12 recovery ratio within the established range (100 ± 20%) [67]. Subsequently, B12 was quantified by HPLC with no interferences with the analytical method.
