Resonance Raman and Multivariate Curve Resolution-Alternating Least Squares (MCR-ALS) Analysis of Biological Polyenes in Shells: A Nondestructive Approach for Species and Habitat Identification

Biogenic minerals such as shells contain trace amounts of biological polyenes derived from carotenoids, which have attracted considerable attention in biology, earth science, environmental science, and materials science. Moreover, the recent issue of fraudulent labeling of edible shellfish origins underscores the need for reliable, nondestructive methods to identify species and habitats. Here, we applied resonance Raman spectroscopy to nondestructively detect carotenoid-derived polyenes (biological polyenes) in various shells differing in species and growing environments. We focused on the peak positions and full width at half-maximum (FWHM) of the bands at around 1500 and 1120 cm(-1), assigned to CC and C-C stretching vibrations, respectively. Single-point Raman measurements revealed that both the peak positions and the FWHM values varied among samples, indicating that multiple types of biological polyenes are present within each shell and that their distribution depends on species and habitats. To further resolve these spectral features, we performed multipoint (Raman imaging) measurements and applied multivariate curve resolution-alternating least squares (MCR-ALS) analysis to extract pure component spectra of the biological polyenes. As a result, multiple distinct polyene species were identified in each shell. Notably, the observed bands at around 1120-1100 cm(-1) differed from those typical of carotenoids, suggesting that shell-derived polyenes vary according to both species and environmental conditions. For a clearer comparison, we categorized these peak positions into four distinct groups. Collectively, our results demonstrate, for the first time, that the types and numbers of biological polyenes differ among shellfish species and habitats without the need for extraction or isolation. This finding implies that shellfish biosynthesize diverse polyenes distinct from carotenoids in response to environmental factors, such as feeding conditions. The combined use of resonance Raman spectroscopy and MCR-ALS thus provides an effective approach for identifying and characterizing biogenic minerals, such as shells.