Editorial Feature

What is Diffuse Reflectance Spectroscopy?

Diffuse reflectance spectroscopy is a type of absorption spectroscopy where, rather than measuring the transmitted beam through the sample, the light reflected from the sample is measured instead. It is a common method for measuring opaque samples that absorb too strongly to be measured in transmission.

diffuse reflectance spectroscopy

Image Credit: Rabbitmindphoto/Shutterstock.com

The information in a diffuse reflectance spectrum can be used for quantitative and qualitative analysis of samples.1 Typically measured using visible or near-infrared light, the resulting spectrum contains information on the electronic or potentially vibrational structure of the sample being probed.

A reflectance measurement is relatively straightforward as it involves sample illumination using a light source and then a detector that collects the scattered light. Instruments will typically contain dispersive gratings so several wavelengths can be detected simultaneously. There is no need for moving parts as in a monochromator system.

Many devices use fiber optic bundles to send and detect the light, particularly for more portable applications or for heritage science where it may be desirable to record spatially resolved spectra over a large painting area.

Instruments may also need several additional optics to separate unwanted scattering or include an integrating sphere in the probe to ensure that as much of the reflected light is collected as possible.

Some advantages of measuring spectra in reflective geometries include improved signal-to-noise and sensitivity versus transmission measurements, but this comes at the cost of more difficult-to-record and complex spectra to understand.1  

Kubelka-Munk theory is a common approach to understanding the scattering and absorbance characteristics of the sample being measured. However, one of the challenges of these measurements is that the incident angle of reflection of the measurement will ultimately influence the final spectrum obtained.

Despite the challenges with measurement interpretation and the need for care when performing experimental measurements, diffuse reflectance spectroscopy is now widely used in many industrial and research environments as it allows for direct measurements of opaque samples that can be challenging for many spectroscopies.

Common Applications of Diffuse Reflectance Spectroscopy

Pharmaceutical industry

One area of application for diffuse reflectance spectroscopy is in the pharmaceutical industry. Diffuse reflectance spectroscopy is commonly used to analyze pharmaceutical compounds and active pharmaceutical ingredients when pressed into the delivery matrix.2

For such applications, near-infrared radiation is a common choice. Many compounds of interest have a series of structured bands in the spectrum that can be used for easier sample identification.

Diffuse reflectance spectroscopy offers a rapid and non-destructive testing technique that requires little additional sample preparation and can be incorporated into online testing environments for an online process analysis as part of a process analytical technology.3

Typical analytical chemistry approaches such as chemometrics methods can be used to analyze datasets of the diffuse reflectance spectra of more complex mixtures and provide concentration and species identification information as part of quality control measures in manufacturing.

Portable measurements: Heritage science applications

The non-destructive nature of diffuse reflectance spectroscopy has also made it a widely used tool in heritage science.4

Absorption and reflectance spectroscopies can be used to identify pigments and reveal hidden murals and paintings that have been covered over by another piece of artwork.

Many heritage science applications require field measurements of artwork and architecture. This means there is a need for portable diffuse reflectance spectrometers that are sufficiently robust, where components do not become misaligned in the field and do not need constant recalibration. For diffuse reflectance spectroscopy, this often means using instruments that use fiber optic bundles to transmit and detect the reflected light.  

The use of monolithic optical mounts and brighter compact light sources have helped make the performance of portable reflectance spectrometers nearly as good as bulky benchtop instruments.5 Advances in automated analysis and application of algorithms such as the Kramer-Kroning relations can help with rapid extraction of spectroscopic data in the field.

Diffuse reflectance spectroscopy has been used as a tool for the diagnosis of cancers and other diseases in the medical sciences.6

The development of more portable, handheld instruments has opened up the possibility for new point-of-care devices that can be used to diagnose patients without needing surgery and biopsies.

The chemical information that can be recovered from the diffuse reflectance spectrum provides information on various biomarkers that can be used to distinguish malignant and benign tumors and make a rapid clinical diagnosis.

There are now a number of cancers that can be diagnosed with diffuse reflectance spectroscopy measurements and with further improvements in the automated analysis of such spectra, diffuse reflectance spectroscopy will likely become part of a suite of spectroscopic techniques, including Raman and infrared measurements, that help reduce diagnosis times and improve clinical accuracy.7

References and Further Reading

  1. Blitz, J. P. (1998) Diffuse Reflectance Spectroscopy. Wiley.  https://www.wiley.com/en-us/Modern+Techniques+in+Applied+Molecular+Spectroscopy-p-9780471123590
  2. Li, P., Du, G., Cai, W., & Shao, X. (2012) Rapid and nondestructive analysis of pharmaceutical products using near-infrared diffuse reflectance spectroscopy. Journal of Pharmaceutical and Biomedical Analysis, 70, 288–294. https://doi.org/10.1016/j.jpba.2012.07.013
  3. Lopes, A., Costa, P. F., Alves, T. P., & Menezes, J. C. (2004) Chemometrics in bioprocess engineering : process analytical technology ( PAT ) applications. Chemometrics and Intelligent Laboratory Systems, 74, 269–275. https://doi.org/10.1016/j.chemolab.2004.07.006
  4. Dal, J. S. A., & Fontana, F. R. (2020) Reflectance imaging spectroscopy in heritage science. La Rivista Del Nuovo Cimento, 43(10), 515–566. https://doi.org/10.1007/s40766-020-00011-6
  5. Arrizabalaga, I., Gómez-laserna, O., Aramendia, J., Arana, G., & Madariaga, J. M. (2014) Applicability of a Diffuse Reflectance Infrared Fourier Transform handheld spectrometer to perform in situ analyses on Cultural Heritage materials. Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 129, 259–267. https://doi.org/10.1016/j.saa.2014.03.096
  6. Evers, D. J., Nachabe, R., Peeters, M. V., Hage, J. van der, Oldenburg, H. S., Rutgers, E. J., Lucassen, G. W., Henddriks, B. H. W., Wesseling, J., & Ruers, T. J. M. (2013) Diffuse reflectance spectroscopy : towards clinical application in breast cancer. Breast Cancer Res Treat, 137, 155–165. https://doi.org/10.1007/s10549-012-2350-8
  7. Akter, S., Hossain, M. G., Nishidate, I., Hazama, H., & Awazu, K. (2018) Medical applications of reflectance spectroscopy in the diffusive and sub-diffusive regimes. Journal of Near Infrared Spectroscopy26(6), 337-350. https://doi.org/10.1177/096703351880663

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Rebecca Ingle, Ph.D

Written by

Rebecca Ingle, Ph.D

Dr. Rebecca Ingle is a researcher in the field of ultrafast spectroscopy, where she specializes in using X-ray and optical spectroscopies to track precisely what happens during light-triggered chemical reactions.


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