HPLC method development/validation and skin diffusion study of caffeine, methyl paraben and butyl paraben as...

The focus of this research was to develop and validate a suitable HPLC method, which allows simultaneous determination of three proposed skin model penetrants to investigate the percutaneous diffusion behavior of their combination: caffeine, methyl paraben and butyl paraben. These penetrants were selected because they represent a wide range of lipophilicities. This model highlights the effect of combining penetrants of different molecular properties on their diffusion behavior through skin. The proposed method employed a gradient system that was systematically optimized for separation and quantification of the penetrants. The effect of the stationary phase (C18, C4 and cyano (CN)) was assessed with CN proven to be superior in terms of peak shape, retentivity and dynamic linear range. Significant differences in retention time, peak broadening, and quantifiability between different stationary phases could be demonstrated. The method was validated as per ICH guidelines Q2 (R1) with a satisfactory outcome. The method was successfully applied for real diffusion experiments, and revealed notable differences between the individual penetrants and their ternary mixture on transdermal permeation. The method could potentially be extended to determine these analytes in other related skin permeation investigations.