Tanja Maria Greve

ATR-FTIR, FT-NIR and near-FT-Raman spectroscopic studies of molecular composition in human skin in vivo and pig ear skin in vitro

ATR-FTIR, FT-NIR and near-FT-Raman spectroscopy were used to characterize the molecular composition of human skin in vivo and pig ear skin in vitro. Due to different measurement depths the spectroscopic techniques reveal the characteristics of different layers of the skin. Tape stripping was used with the ATR-FTIR technique. Spectral differences concerning lipid content and conformation, protein secondary structure or content of water were found with respect to both gender and species (i.e. human versus pig ear) at all measured skin depths. New assignments of so far unassigned lipid and protein peaks in the FT-NIR and ATR-FTIR spectra of skin were made. PCA and PLS models were used to investigate the division of the recorded spectra into groups. With respect to classification of male and female subjects, the PLS discriminant analysis provided a classification accuracy of 64-93% based on the ATR-FTIR spectra and 83-89% based on the Raman spectra. With respect to classification of human skin in vivo and pig ear skin in vitro, the PLS discriminant analysis provided a classification accuracy of 75-100% based on the Raman spectra and 100% based on the ATR-FTIR spectra.

Imaging of Distribution of Topically Applied Drug Molecules in Mouse Skin by Combination of Time-of-Flight Secondary Ion Mass Spectrometry and Scanning Electron Microscopy

In the development of topical drugs intended for local effects in the skin, one of the major challenges is to achieve drug penetration through the external barrier of the skin, stratum corneum, and secure exposure to the viable skin layers. Mass spectrometric imaging offers an opportunity to study drug penetration in a variety of skin models by mapping the spatial distribution in different skin layers after topical application of the drug. In this study, we used time-of-flight secondary ion mass spectrometry (TOF-SIMS) and scanning electron microscopy (SEM) to image the distribution of three drug molecules in skin tissue cross sections of inflamed mouse ear. The three compounds, roflumilast, tofacitinib, and ruxolitinib, were topically administered to the mouse ears, which were subsequently cryosectioned and thawed for the analyses. The results reveal that the combination of TOF-SIMS and SEM was beneficial for interpretation of drug distribution. SEM identified the different skin layers, while spatial distributions of all three compounds could be visualized by TOF-SIMS, showing that the drug was primarily distributed into, or on the top of, the stratum corneum. Imaging of endogenous skin components like cholesterol, phospholipids, ceramides, and free fatty acids showed distributions in good agreement with the literature. One limitation of the TOF-SIMS method is sensitivity, typically allowing for analysis in the millimolar range rather than the pharmacologically relevant micromolar range. However, the data presented demonstrate the potential of the technique for studying the penetration of drugs with different physicochemical properties in skin.

Penetration mechanism of dimethyl sulfoxide in human and pig ear skin: An ATR–FTIR and near-FT Raman spectroscopic in vivo and in vitro study

The penetration mechanism of dimethyl sulfoxide (DMSO) in human skin in vivo and in vitro and pig ear skin in vitro was studied using attenuated total reflectance (ATR) Fourier transform (FT) infrared (IR) and near-FT-Raman spectroscopy. The results showed changes in the conformation of the skin keratins from an α-helical to a β-sheet conformation. These changes were proved to depend on the concentration of free water in the sample as DMSO tended to bind to free water before the protein-bound water was replaced and the protein conformational changes were induced. The induced conformational changes were shown to be completely reversible as the proteins are returned to their original state within 20 h after the treatment with DMSO. The penetration depth of DMSO was shown to depend on the time of exposure - however, after only 15 min DMSO has penetrated the stratum corneum, which is the skin barrier.

Disease quantification in dermatology: in vivo near-infrared spectroscopy measures correlate strongly with the clinical assessment of psoriasis severity

Abstract. Accurate documentation of disease severity is a prerequisite for clinical research and the practice of evidence-based medicine. The quantification of skin diseases such as psoriasis currently relies heavily on clinical scores. Although these clinical scoring methods are well established and very useful in quantifying disease severity, they require an extensive clinical experience and carry a risk of subjectivity. We explore the opportunity to use in vivo near-infrared (NIR) spectra as an objective and noninvasive method for local disease severity assessment in 31 psoriasis patients in whom selected plaques were scored clinically. A partial least squares (PLS) regression model was used to analyze and predict the severity scores on the NIR spectra of psoriatic and uninvolved skin. The correlation between predicted and clinically assigned scores was R=0.94 (RMSE=0.96), suggesting that in vivo NIR provides accurate clinical quantification of psoriatic plaques. Hence, NIR may be a practical solution to clinical severity assessment of psoriasis, providing a continuous, linear, numerical value of severity.