Researchers refine Raman spectroscopy to characterize melanin more reliably
ICFO researchers have developed a strategy that overcomes the challenges associated with Raman spectroscopy when characterizing the two main forms of melanin. The methodology, published in Microchemical Journal, reduces signal distortions and facilitates comparisons between differently pigmentated samples and across varying experimental conditions. These results could guide future research on melanoma disease.
Melanin, the pigment that gives color to our skin, hair and eyes, is also believed to play a role in melanoma disease –the most dangerous form of skin cancer. To date, two main forms of melanin have been identified: eumelanin, which is photoprotective and antioxidant, and pheomelanin, which is phototoxic and prooxidant. Being able to correctly identify and quantify them could advance melanoma research, either by deepening our understanding of how the cancer works or by providing insights into effective treatment strategies. However, gold-standard methods for melanin characterization are destructive and require extensive sample preparation. Therefore, there is growing interest in developing non-destructive optical approaches for melanin analysis.
Raman spectroscopy is one of the non-destructive optical approaches currently being explored for melanin analysis, as it provides molecular fingerprints of samples when illuminated by a laser. However, melanin analysis by Raman spectroscopy remains challenging due to the strong autofluorescence background and instrumental artefacts that can distort the measured spectra. Now, ICFO researchers, José Javier Ruiz and Dr. Pablo Loza-Álvarez, Chief of the SLN Facility, together with Ismael Galván from the National Museum of Natural Sciences (CSIC), have developed a strategy to overcome the challenges it presents. Published in Microchemical Journal, the strategy –which was tested using hairs and feathers with different pigmentation– has shown that Raman imaging can reliably retrieve and characterize melanin information from intact biological samples.
There are two main difficulties that the proposal overcomes. First, it corrects a common distortion that complicates the interpretation of data, whose instrumental origin had not been clearly identified and corrected until now. “Several previous studies had observed a sinusoidal interference, which led to divergent results,” explains José Javier Ruiz, first author of the article. “In our study, we describe how to characterize and correct it.”
Secondly, it addresses the broad and intense background signal caused by autofluorescence, which often masks the weaker Raman signals of interest. Rather than treating this autofluorescence solely as an unwanted interference, the proposal also takes advantage of it to provide complementary information about sample pigmentation.
“We anticipate that these advances may support future Raman and in general optically-based studies of different biological samples containing melanin, including research into melanocytic tumors”, says Dr. Pablo Loza-Álvarez, lead researcher of the study. “Improving the reliability and comparability of melanin spectral analysis, and making it more robust across different imaging platforms, could help to better understand how melanin composition and organization relate to different biological and pathological conditions.”
Reference:
José Javier Ruiz, Ismael Galván, Pablo Loza-Alvarez, Correction of Rayleigh filter ripple and fluorescence background to enable reliable Raman analysis of melanin, Microchemical Journal, 225, 2026, 118178.
DOI: https://doi.org/10.1016/j.microc.2026.118178
Acknowledgements:
This research was funded by CEX2024-001490-S [MICIU/AEI/ 10.13039/501100011033], Fundació Cellex, Fundació Mir-Puig, and Generalitat de Catalunya through CERCA, and COST Action (CA23125). The SLN facility corresponds to a “Grup reconegut” 2021 SGR 01456- Departament de Recerca i Universitats de la Generalitat de Catalunya.