Quantitative detection of caffeine in human skin by confocal Raman spectroscopy – A systematic in vitro validation study

• We introduce a procedure to achieve homogeneous drug concentrations in human skin.
• We detect the drug by Raman spectroscopic depth profiles in excised human skin.
• We validate Raman spectroscopic measurements by HPLC quantification.
• We evaluate skin derived Raman peaks as internal standard for peak correlation.
• We establish a linear dependency of Raman signal and drug amount over depth.

For rational development and evaluation of dermal drug delivery, the knowledge of rate and extent of substance penetration into the human skin is essential. However, current analytical procedures are destructive, labor intense and lack a defined spatial resolution. In this context, confocal Raman microscopy bares the potential to overcome current limitations in drug depth profiling. Confocal Raman microscopy already proved its suitability for the acquisition of qualitative penetration profiles, but a comprehensive investigation regarding its suitability for quantitative measurements inside the human skin is still missing.In this work, we present a systematic validation study to deploy confocal Raman microscopy for quantitative drug depth profiling in human skin. After we validated our Raman microscopic setup, we successfully established an experimental procedure that allows correlating the Raman signal of a model drug with its controlled concentration in human skin. To overcome current drawbacks in drug depth profiling, we evaluated different modes of peak correlation for quantitative Raman measurements and offer a suitable operating procedure for quantitative drug depth profiling in human skin.In conclusion, we successfully demonstrate the potential of confocal Raman microscopy for quantitative drug depth profiling in human skin as valuable alternative to destructive state-of-the-art techniques.

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