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Issue
32— January 2003 |
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Titel |
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Authors |
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Article |
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Analysis
of the stratum corneum by attenuated total reflection spectroscopy using
a novel mid-infrared fibre probe |
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H.
Michael Heise, Lukas Küpper, Wolfgang Pittermann, and Manfred Kietzmann |
Summary
Infrared attenuated total reflection (ATR) spectroscopy is an established method
for stratum corneum analysis. The potential for dermatology studies is increased
by the development of a flexible fibre-optic probe made from infrared-transparent
polycrystalline silver halide material, which eases the epidermal surface characterisation
significantly. As a substitute for human in-vivo tests, the isolated perfused
bovine udder skin (BUS-model) has been proposed. The chemistry of human skin and
of bovine udder skin is compared on the basis of their infrared spectra. For depth
profiling, subsequent stripping of corneocyte layers by adhesive tape application
was carried out. An example of a topical cream application and the measurement
of the penetration profile of the cream compounds in the stratum corneum by the
fibre-probe ATR-measurements and subsequent tape stripping are presented.
For studies of the upper horny skin layer, attenuated total reflection (ATR) infrared
spectroscopy has often been used, allowing a shallow micrometer probing depth
for the surface measurement. The range of applications is increased by the development
of a fiber-optic probe made from polycrystalline infrared-transparent silver halide
material with ductile and non-toxic characteristics, (1) which provides more flexibility
for the epidermal surface characterisation than available with conventional accessories.
Largest transmittance of the fibres is observed particularly within the information-rich
infrared fingerprint region (about 1500 - 600 cm-1) (2). Our aim was to study
the chemistry of the human horny layer and that of the upper bovine udder skin,
and a comparison was made possible on the grounds of their ATR-spectra. Further
potential of the measurement technique in combination with adhesive-tape stripping
for penetration studies of topically applied active substances is illustrated.
The in-vitro model of isolated perfused bovine udder skin (BUS-model) has been
proposed as a substitute for human in vivo tests (3). Due to the continuous perfusion,
the horny layer demonstrates active barrier and reservoir functions. The in vitro
BUS model is widely used in dermatological and cosmetic research as well and exhibits
hair follicles and sebaceous glands, providing the corneal compartment with sebum
similar to the human in vivo situation. Histological studies also prove the similarity
of the BUS-model to human skin.
In Figure 1, the measurement instrument, including the attachment of the
fibre-probe, is shown.
The spectrometer was a Fourier-transform infrared (FTIR)-spectrometer. The set-up,
with mirror optics, fibre-probe and fibre-detector coupling, was a development
of the Institute of Spectrochemistry and Applied Spectroscopy and Infrared Fiber
Sensors, which includes a semiconductive mercury-cadmium-telluride (MCT)-detector.
The silver halide fibre-optic probe consisted of a shaft with one fibre for transmitting
and a second one for receiving infrared radiation from the ATR-measuring head.
The measurement principle, with the evanescent electrical radiation field outside
the optically dense fibre, which is either air or the sample in contact with the
fibre for the respective background or sample measurement, is illustrated in Figure
1 b. The transmitting and receiving fibres were 2 m in length (either of square
cross-section of 750 µm x 750 µm or of circular cross-section with
a diameter of 700 µm; optical numerical aperture of 0.5). The shaft length
was 20 cm (12 mm in diameter), and one prototype also had means for attaching
detachable short U-shaped silver halide fibre pieces for ATR measurements (see
Figure 1 c).
An example for the spectral measurements of bovine udder and human forearm skin
using the fibre-optic ATR-probe is provided in Figure 2.
Skin areas of less than 1 mm² were measured with a probe made from a silver
halide fibre of square cross-section. Repeat skin stripping by application of
adhesive tape was also performed, similar to the technique described recently
(4). Besides spectra measured from the skin surface, a spectrum of the lipids
within the udder skin and of sebum from a human male forehead are also shown (Figure
2 a and 2 b, respectively). Differences in the spectra are due to keratin differences
(see the strong amide I bands at 1650 cm-1, mainly C=O stretching vibration, and
the most striking features in the C-O and C-C-stretching region between 1150 and
1000 cm-1) and the amount of lipids observed. For the latter, the absorption band
of the ester C=O stretching at 1740 cm-1 and an overlapped band at about 1460
cm-1 from CH2-deformation vibrations are characteristic. There is an astonishing
similarity in both lipid spectra, giving evidence to a similar chemistry, apart
from the absorption band at 1710 cm-1, which is found in the human sebum spectrum
and can be assigned to a free fatty acid component.
In Figure 3 the exemplary results from experiments with the application
of a lamellar cream applied to udder skin are shown.
In Figure 3 a, the ATR-spectrum of a dry film of the cosmetic cream is also given,
which eases the interpretation of the in-situ measurements that were carried out
after subsequent tape-stripping. The disappearance of cream constituents is obvious
after 10 tape-strippings. Complementary information is obtained from the measurements
on the adhesive tapes used (Figure 3 b), which can be exploited also for a quantitative
determination of the stripped corneal material and cream constituents.
Conclusion
The non-invasive infrared spectroscopic measurement technique can be applied to
human studies as well as in vitro studies using skin models with fully functional
horny layers and natural sebum excretion. The technique, based on probes made
from fibres, in particular of square cross-section, is very promising, since it
opens the field for new medical and cosmetic applications that were not possible
with conventional ATR-crystals and bulky sampling compartment-based accessories.
Acknowledgements:
H.M. Heise and L. Küpper from the Institute of Spectrochemistry and Applied
Spectroscopy acknowledge gratefully the financial support by the Ministerium für
Schule, Wissenschaft und Forschung des Landes Nordrhein-Westfalen and the Bundesministerium
für Bildung und Forschung.
References
1. L. Küpper, H.M. Heise, L.N. Butvina, Novel developments in mid-IR fiber
optic spectroscopy for analytical applications, J. Mol. Struct. 563/564, 173-181
(2001)
2. L. Küpper, H.M. Heise, W. Pittermann, L.N. Butvina, New tool for epidermal
and cosmetic formulation studies by attenuated total reflection spectroscopy using
a flexible mid-infrared fiber probe, Fresenius J. Anal. Chem. 371, 753-757 (2001)
3. Th. Förster, W. Pittermann, M. Schmitt, M. Kietzmann, Skin penetration
properties of cosmetic formulations using a perfused bovine udder model, J. Cosmet.
Sci. 50, 147-157 (1999)
4. C. Laugel; C. Do Nascimento; D. Ferrier; J.P. Marty; A Baillet, Contribution
of ATR/FT-IR spectroscopy for studying the in vivo behavior of octylmethoxycinnamate
(OMC) after topical application, Appl. Spectrosc. 55, 1173-1180 (2001)
Author
Dr. H. Michael Heise
Dr. H. Michael Heise
Institut für Spektrochemie und Angewandte Spektroskopie
Bunsen-Kirchhoff-Str. 11
D-44139 Dortmund
Tel.: +49 231 1392215
Fax: +49 231 1392120
E-mail: heise@isas-dortmund.de
Dipl.-Chem. Dr. rer. nat. H. Michael Heise took up his current appointment at
the Institute of Spectrochemistry and Applied Spectroscopy at the University of
Dortmund in 1982. Current research interests are in near- and mid-infrared spectroscopy
and chemometrics, especially with analytical applications to clinical chemistry
and medical diagnostics. After obtaining his PhD in 1976 he carried out postdoctoral
work at the University of Western Australia in Perth, at the Federal Biological
Research Centre and the Physikalisch-Technische Bundesanstalt in Braunschweig
(Germany). He is the author and coauthor of more than 130 publications, books,
and book chapters.
This article was coautored by Dr. Lukas Küpper, Infrared Fiber Sensors (IFS),
Aachen, Germany, Dr. Wolfgang Pittermann, Henkel KGaA, Düsseldorf, Germany
and Prof. Dr. M. Kietzmann, Institute for Pharmacology, Toxicology and Pharmacy,
Veterinary School Hannover, Hannover, Germany.
