Skin Research and Technology zyxwvutsrqpon 1997; 3: 147-154 Printed in Denmark. All rights reserved Cogtlrifht zyxw 0 Munksfaard zyx 1997 Skin Research and zy Technology ISSN 0909-752X ReviewArticle Applications of zyxw Raman spectroscopy to skin research Emma E. Lawsonl, Howell G. M. Edwards2, Adrian C. Williams' and Brian W. Barry' 'Postgraduate Studies in Pharmaceutical Technology, The School zyxwvuts of Pharmacy and 2Chemist y and Chemical Technology, University of Bradford, Bradford, UK zyxwvut Backgroundlaims: zyxwvutsrqp Raman spectroscopy has been used for a range of biomedical applications: the study of normal and dis- eased tissues, and the interaction of chemical agents with tissues, implants and even single cells. The object here was to review the extent to which the Raman spectroscopic technique has been applied to skin research, considering the implications of different instrumentation, comparing animal and human skin, healthy and diseased skin and in vivo and in vitro sampling. Conclusions: Raman spectroscopy is a versatile and non-de- structive technique for the study of skin. Key words: Raman spectroscopy - skin - biomedical - instru- mentation - animal - human - in vivo, in vitro 0 Munksgaard, 1997 Accepted for publication 25 November 1996 AMAN SPECTROSCOPY has been used in a variety of R biomedical applications, including the study of normal and diseased human tissues (1-5), single cells (6) and implants (7), the presence of foreign inclusions following implantation (8,9) and the interaction of cer- tain chemical agents with tissues (10-12). In general, biomedical studies using Raman spectroscopy only became well established with the introduction of near- infrared region Fourier transform (NIR FT) Raman spectroscopy in the mid- to late 1980's. This advance- ment in technology marked the beginning of an up- surge in tissue studies, which had previously been hampered by sample fluorescence, with noted inno- vators in the biomedical field being Yu et al. (13) and Ozaki et al. (14). Charge coupled diode (CCD) Raman spectrometers operating in the NIR (780-900 nm) offer faster scan times than FT-Raman spectroscopy, al- though the use of CCD systems is still quite uncom- mon in the field of biomedical applications. The ad- vantages of coupling optical fibre devices to Raman spectrometers for facilitating in vivo studies are gen- erally acknowledged as showing potential in the sphere of diagnostics although this area of research is still quite new. Raman Spectroscopy Raman spectroscopy is a spectroscopic technique that provides qualitative and quantitative evaluation of the structures and transformation of materials at the molecular level in terms of molecular normal vi- brational frequencies. In Raman spectroscopy the sample, which may be liquid, solid or gaseous, is irradiated with an intense beam of monochromatic radiation of wavenumber, uo. Most of this radiation is transmitted by the sample with a small part being scattered both elastically, with a wavenumber equal to that of the incident radiation, and inelastically, with wavenumbers that differ from those of the incident radiation. Most of the scatter is elastic and is known as Rayleigh scattering. A very small part of the scattered radiation (lop5 of the inci- dent radiation intensity) is inelastically scattered with wavenumbers of uO+um, where urn is a characteristic vibrational wavenumber of the molecular species that is undergoing excitation. This is known as Raman scattering, occurring when a photon (uo) of the inci- dent beam collides with a molecule causing one of the normal modes of vibration (u,) to increase its vi- brational energy at the expense of the incident pho- ton. The incident photon is annihilated and a new photon (uo+um) is scattered. The uo-um (Stokes) and uo+u, (anti-Stokes) scat- tered radiation components are observed at wave- numbers lower and higher than those of the Rayleigh scattering, respectively (Fig. 1). Depending on whether the molecule was occupying the ground state energy level or the first excited state energy level, the scattered radiation will be Stokes or anti-Stokes, re- spectively (Fig. 1). The two main types of Raman spectrometers im- plement either dispersive or Fourier transform (FT) techniques and both comprise four basic instrument components: 1) a radiation source (usually a laser), 2) 147