Insights in the biochemical composition of skin as investigated by micro infrared spectroscopic imaging

Abstract

The conventional methods for biochemical and histophatological tissue characterisation necessitate complex and “time consuming” sample manipulations and the results are rarely quantifiable. Micro infrared imaging is a relatively new approach for the examination of tissue sections. In contrast to standard light microscopy based procedures, the IR approach requires neither sample staining nor complicated fixation. The contrast in infrared images is solely derived from differences in the spectral and spatial heterogeneity of the biochemical components. In this study the morphological organisation and biochemical composition of “Göttingen” pig skin is analysed. The upper layer of the skin, the so called stratum corneum, is composed of corneocytes (terminally differentiated cells) imbedded in a lipid matrix. Based on the integral intensity of specific bio-marker absorption bands for the lipid as well as for the protein fraction (cells), infrared images were generated (functional group imaging). These infrared images represent the biochemical distribution of the analysed fraction within the sample. The characteristic bio-marker bands for the lipids are: the asymmetric (∼2920 cm⁻¹) and symmetric methylene (∼2851 cm⁻¹) stretching absorptions, and the carbonyl (∼1740 cm⁻¹) vibration. The protein fraction is imaged by using the integral intensity of the asymmetric methyl vibration (∼2957 cm⁻¹) as well as by the amide I (∼1655 cm⁻¹), amide II (∼1545 cm⁻¹) and amide B (∼3080 cm⁻¹) absorptions. Evidence is given, that the upper stratum corneum is characterised by large areas of lipids and aggregated corneocytes. Phase separation is also observed for the deeper stratum corneum layers. However, the detected domains are much smaller. This demonstrates that the stratum corneum is not so regularly organised as described in the simplified “bricks and mortar” model. The different layers of the skin (stratum corneum, epidermis and dermis) are easily made visible by using the presented imaging technique and are fairly different concerning its biochemical composition. Micro infrared imaging is also used for analysing the penetration and distribution of a sun cream lotion in the skin. Depending on the used formulation, the distribution in the tissue can be quite different. The presented example demonstrates how this technique can be used for the development of topical applied formulations. The results in this report suggest that micro infrared spectroscopic imaging may become a new and promising investigation tool in bio-medicinal research and will provide a wealth of information on tissues that is not readily accessible by other physical and biochemical techniques.