2.4. Transdermal Delivery In Vitro Using Franz-Type Diffusion Cells

A porcine skin membrane having the dimensions 2.5 × 2.5 cm was purchased (Micropig Franz Cell Membrane, Apures, Pyeongtaek-si, Korea). Before the experiment, the skin membrane was immersed in PBS (Phosphate buffered saline pH 7.4, Gibco, Waltham, MA, USA) and incubated for 2 h. In the following experiment, the surface of the stratum corneum was wiped with a paper tissue to get rid of the extra PBS. For the application of microneedles, the array was inserted into the skin under gentle thumb pressure. The reservoir was filled with PBS. The diffusion cells were tightly assembled with clamps and placed in an incubator at 37 °C and 50% relative humidity (RH). To quantify the target molecule in the stratum corneum, epidermis, and dermis, respectively, the porcine skin was tape-stripped three times and heated at 90 °C for 50 s. After separation of the epidermis and dermis, each compartment was homogenized with PBS and centrifuged. The clear supernatants were recovered and analyzed using photoluminescence spectroscopy (Varioskan™ LUX, Thermofisher, Waltham, MA, USA) to quantify the FITC and an Ascorbic Acid Assay Kit (L-Ascorbate, K-ASCO, Megazyme, Bray, UK) to quantify the vitamin C. The amount of retinol was analyzed by HPLC. Detailed analysis followed the previous literature [18].

For the transdermal delivery of ascorbic acid and FITC, ascorbic acid and FITC were prepared with concentration of 25% and 50,000 ng/mL, respectively. A 25% solution of vitamin C was titrated to pH 3.2 with KOH.