Abstract
Pharmaceutical products usually comprise of active ingredients and a number of excipients. The amount of the actual drug can be very small and hence there is a need for highly sensitive and specific techniques for quantitative and qualitative analysis. Surface-enhanced Raman spectroscopy (SERS) is introduced as a particularly promising tool in this context. For example, it can be employed for analyzing pharmaceutical products, but it can also help to better understand the interactions between pharmaceutically active compounds with pathogenic germs and biological organisms and, thereby, it may contribute to the development of target-specific treatments, e.g. for the currently spreading corona virus disease, COVID-19.
Introduction
Over the past decades, Raman spectroscopy has been established as a versatile tool in the pharmaceutical sector. It can not only be used for the qualitative and quantitative analysis of products, but can also be applied as a means of Process Analytical Technology (PAT). However, as it is based on inelastic scattering of light, Raman spectroscopy signals are typically rather weak. To some extent, this disadvantage can be compensated by using highly sensitive detectors and high-power lasers. However, when the analyte to be determined is present in a very small amount, and/or exhibits a low scattering cross section, and/or is susceptible to photo-degradation, the “More Power!” approach is not the method of choice. In such a situation, other techniques to enhance the signal are required.
The specificity and sensitivity of Raman spectroscopy can be improved by using a laser wavelength that is in electronic resonance with the species of interest. This approach is known as resonance Raman scattering. Its disadvantage is, however, that it requires a more flexible and hence more expensive laser source. Surface-enhanced Raman scattering (SERS) is a suitable alternative. In SERS, the sample is brought in contact with a plasmonic substrate, typically a metal such as silver or gold. The SERS substrate can be a roughened metal surface, metal nanoparticles disposed on a glass or polymer surface, or a colloidal solution. The general effect that such a metal surface can enhance the Raman signals was first reported in 1973.1 It did not take long to realize the great potential of SERS in the biochemical and pharmaceutical sectors, see for example the review articles.2-4
Currently, there are two mechanisms that are believed to yield the signal enhancement: the plasmonic effect and the chemical effect. When the electromagnetic wave hits the surface and is in resonance, it can excite localized plasmons in a sense that the free electrons in the metal oscillate together with the electric field. This is illustrated in Figure 1. As a consequence, the electric field in the close vicinity of the surface is enhanced and hence there is a significant increase in the Raman signal intensity if the molecule of interest is in the range of this field enhancement. The signal levels can increase by several orders of magnitude allowing even single-molecule detection in specific cases.5 On the other hand, the chemical effect is based on a charge transfer between the surface and the molecule. For this transfer to happen, the molecule needs to be adsorbed at the surface.
Overall, enhancement factors of up to about 1010 have been reported, highlighting the great sensitivity that can be achieved by SERS. On the other hand, SERS has a disadvantage: its limited reproducibility. As described above, the surface enhancement is a highly localized effect taking place at the interface between the substrate and the sample. The substrate surface is nano-structured and the target molecules often have multiple sites capable of interacting with the substrate.
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This results in an adsorption behavior that is difficult to control or to predict. As a consequence, repeating a SERS experiment may yield slightly different spectra. The same is true for the comparison between conventional Raman spectra and SERS spectra. However, it should be pointed out that this mainly affects the quantitative analysis; qualitative analysis is possible with high specifi city and sensitivity.
In order to illustrate the difference between Raman and SERS, caffeine has been chosen as an example. The intensity-normalized Raman and SERS spectra are displayed in Figure 2. The spectra are calculated assuming Gaussian peak shapes and utilizing the peak position and intensity data of an experimental study.6 The spectra show that virtually all of the peaks present in the Raman spectrum can also be found in the SERS spectrum. The SERS data reveal a number of additional peaks and a significantly different spectral shape in terms of the relative intensities across the spectrum. This is due to the local charge transfer and plasmonic enhancement.
SERS in the Pharmaceutical Sector
There are a number of highly interesting applications of SERS in the context of pharmaceuticals. For example, SERS can be used to detect pharmaceutically active ingredients in drug products. This feature can be utilized to evaluate the final product quality or intermediates during the production process. A great advantage of the SERS method in this context is its high sensitivity as it allows assessing the presence or absence of even tiny amounts of certain ingredients. Care must be taken, however, when one or more species of interest need to be quantified. This can be difficult due to the limited reproducibility of SERS in some applications for the reasons discussed above. When rapidness is not required, this limitation can be overcome by averaging spectra.
The capability of detecting traces of pharmaceutically active compounds can also be taken advantage of in studies and investigations into the behavior of and exposure to drugs. For example, it is possible to monitor the behavior of active compounds in treated tissue or the human body with the aim to unravel and optimize the drug delivery and biomedical working mechanisms. This can help to better understand the interactions of pharmaceutically active compounds with pathogenic germs and biological organisms and, thereby, it may contribute to the development of target-specific treatments, e.g. for the currently spreading coronavirus disease, COVID-19. Forensics is another area, where the detection of drugs in tissue and body fluids can be of great interest.
Further emerging fields of SERS applications include the detection of drug adulteration and contamination with unwanted species. Adulterated and contaminated pharmaceutical products do not only pose a health risk, but also represent a severe economic threat to the companies producing high quality pharmaceuticals legally. Furthermore, using SERS as a means of Process Analytical Technology is on the rise. For inline applications, however, suitable approaches to ensure contact between a SERS substrate and the sample need to be developed and/or optimized. One option is to seed trace amounts of a colloidal solution with SERS-active nanoparticles into the process fluid. For this to be feasible, the colloid either must not be hazardous or it needs to be removed again afterwards. Another option is to use SERS probes in which the SERS substrate is fixed, e.g. at the end of an optical fiber.
Conclusion
In conclusion, surface-enhanced Raman spectroscopy (SERS) is a rapid, species-specific, and highly sensitive tool. Therefore, it is an interesting analytical method for a variety of applications in the pharmaceutical sector and related fields. It can provide valuable information during the initial screening, development, production, and testing of pharmaceutically active ingredients. And it can be used as a means of product analysis and quality assessment.
References
- M. Fleischmann, P.J. Hendra, A.J. McQuillan, Raman spectra of pyridine adsorbed at a silver electrode, Chemical Physics Letters 26 (1974) 163-166.
- M.T. Alula, Z.T. Mengesha, E. Mwenesongole, Advances in surface-enhanced Raman spectroscopy for analysis of pharmaceuticals: A review, Vibrational Spectroscopy 98 (2018) 50-63.
- T. Frosch, A. Knebl, T. Frosch, Recent advances in nano-photonic techniques for pharmaceutical drug monitoring with emphasis on Raman spectroscopy, Nanophotonics 9 (2020) 19-37.
- J. Cailletaud, C. De Bleye, E. Dumont, P.Y. Sacre, L. Netchacovitch, Y. Gut, M. Boiret, Y.M. Ginot, P. E. Ziemons, Critical review of surface-enhanced Raman spectroscopy applications in the pharmaceutical field, Journal of Pharmaceutical and Biomedical Analysis 147 (2018) 458-472.
- A.B. Zrimsek, N.H. Chiang, M. Mattei, S. Zaleski, M.O. McAnally, C.T. Chapman, A.I. Henry, G.C. Schatz, R.P. Van Duyne, Single-molecule chemistry with surfaceand tip-enhanced Raman spectroscopy, Chemical Reviews 117 (2017) 7583-7613.
- I. Pavel, A. Szeghalmi, D. Moigno, S. Cinta, W. Kiefer, Theoretical and pH dependent surface enhanced Raman spectroscopy study on caffeine, Biopolymers 72 (2003) 25-37.
Author Biography
Prof. Dr. Johannes Kiefer is Chair Professor and Head of the Engineering Thermodynamics department at the University of Bremen, Germany. In addition, he holds a guest professorship of the Erlangen Graduate School in Advanced Optical Technologies (SAOT) at the University Erlangen-Nuremberg, Germany. His research interests are the areas of developing and applying spectroscopic techniques for the characterization of advanced materials and processes.