Transdermal Delivery of Cytochrome C—A 12.4 kDa Protein—Across Intact Skin by Constant–Current Iontophoresis

Abstract

To demonstrate the transdermal iontophoretic delivery of a small (12.4 kDa) protein across intact skin. The iontophoretic transport of Cytochrome c (Cyt c) across porcine ear skin in vitro was investigated and quantified by HPLC. The effect of protein concentration (0.35 and 0.7 mM), current density (0.15, 0.3 or 0.5 mA.cm(-2) applied for 8 h) and competing ions was evaluated. Co-iontophoresis of acetaminophen was employed to quantify the respective contributions of electromigration (EM) and electroosmosis (EO). The data confirmed the transdermal iontophoretic delivery of intact Cyt c. Electromigration was the principal transport mechanism, accounting for approximately 90% of delivery; correlation between EM flux and electrophoretic mobility was consistent with earlier results using small molecules. Modest EO inhibition was observed at 0.5 mA.cm(-2). Cumulative permeation at 0.3 and 0.5 mA.cm(-2) was significantly greater than that at 0.15 mA.cm(-2); fluxes using 0.35 and 0.7 mM Cyt c in the absence of competing ions (J ( tot ) = 182.8 +/- 56.8 and 265.2 +/- 149.1 microg.cm(-2).h(-1), respectively) were statistically equivalent. Formulation in PBS (pH 8.2) confirmed the impact of competing charge carriers; inclusion of approximately 170 mM Na(+) resulted in a 3.9-fold decrease in total flux. Significant amounts ( approximately 0.9 mg.cm(-2) over 8 h) of Cyt c were delivered non-invasively across intact skin by transdermal electrotransport.