Resonance Raman Spectroscopy (RRS) has been established as a quick, non-invasive and highly sensitive method to assess carotenoids in the human skin. RRS data correlate significantly with those of plasma HPLC methods (High-performance liquid chromatography)
Reflectance spectroscopy is yet another non-invasive method for the assessment of carotenoids. The main advantages of reflection spectroscopy-based devices in comparison to Raman devices are the relatively low price, compact size and light weight. Reflection spectroscopy data correlate with RRS and plasma HPLC. However, classical reflection spectroscopy proved to deliver results that were not stable and accurate enough for commercial use since human skin contains hundreds of different chemical substances and consists of multiple distinct layers. Skin tissue is highly complex and inhomogeneous due to the presence of different chromophores (parts of molecules which absorb special wavelengths of visible light and transmit/reflect others), furrows, wrinkles, birthmarks, layered microstructure, etc. Each substance and each layer interacts in a very specific way with light that enters the skin. This makes the light propagation through the skin a very complicated process, which is characterized by reflection, inhomogeneous multiple scattering and (re)absorption.
In order to get more reliable results, spatially resolved reflectance spectroscopy has been developed (SRRS).
ENTER SRRS, aka Spatially resolved reflectance spectroscopy:
In an experimental setup for SRRS, light enters the skin from an illumination chamber which is separated from the detection chamber by a thin wall. Photons are being scattered, reflected and absorbed during their propagation through the medium and finally exit the skin hitting the detectors in the detection chamber. Carotenoids, for instance, are chromophores that absorb blue light. The more carotenoid molecules are present in the tissue, the more photons statistically get absorbed and the less photons ultimately leave the medium and hit the detectors. This way concentrations of carotenoids can be assessed in the skin.
Since spectroscopic devices with a single point irradiation and a single point detection of light are unable to distinguish between the optical signals being influenced by scattering or absorption, spatially resolved spectroscopy was developed. SRRS picks up the light at several positions that have different distances to the light source. This allows distinguishing data where the light has gone farther (and deeper) into the skin from data where the light only propagated over a short distance (and close to the surface). The scattered photons follow banana shaped propagation paths in the skin. Furthermore, in addition to distinction between absorption and scattering, signals from unwanted depths (e.g. blood chromophores like haemoglobin and bilirubin) can be removed from the informative signal.
Monte Carlo simulations and multi-layered skin models are used to compare theoretical results with experimental spectroscopic results. By use of Monte Carlo simulations and multi-layered models of the human skin, it is possible to simulate light propagation, reflectance and absorption in the tissue. It has been shown that Monte Carlo simulations, using appropriate optical and geometrical parameters, are able to produce simulated spectra that agree well with measured spectra. This can be used to better understand measured data and enhance data evaluation and accuracy.
Using Monte Carlo simulations, it is therefore possible to correlate simulated photon propagations in the skin, including entry and exit scenarios, with experimental results and thus relating them to carotenoid concentrations in the tissue.
In order to be able to better assess the influence of chromophores other than carotenoids on the scattering characteristics of the different layers of the skin, LEDs with different colors are used to record propagation characteristics of light with distinct wavelengths in the tissue. These results help to correct and enhance carotenoid measurements and to make them more stable.