Application of a Multi-Technique Approach to the Identification of Mineral Polymorphs in Histological Samples: A Case of Combined Use of SEM/EDS and Micro-Raman Spectroscopy

Abstract

In the last few years, an increasing interest has developed regarding the application of different techniques for the identification of pollutants inside the tissues deriving from patients affected by benign or neoplastic diseases. Particular attention was paid to neoplasia linked to particular exposures, e.g., heavy metals, carbon dusts, silica, asbestos. As regards the last pollutant, a wide body of scientific literature has been collected, considering the severe effects caused by mineral fibers on human health. Optical and electronic microscopies were widely applied to identify the fibers in respiratory and extra-respiratory organs to detect the minerals and to link their presence to an exposure source and to understand their role in cancer development. The main advantage of electron microscopy lies in the possibility of coupling the microscopes with energy dispersive spectrometers and also collecting data on the elemental composition of various inorganic phases. In term of sample preparation and time of analysis, the most utilized microscope technique is Scanning Electron Microscopy with an annexed energy dispersive spectrometer (SEM/EDS), allowing for the morphological and chemical characterization of the observed particles/fibers. Moreover, this technique is envisaged by Italian Law for asbestos identification in air and bulk samples. On the other hand, this technique does not allow a reliable identification of the mineral phase in the case of polymorphs with the same chemical formula but different crystal structures. In this work, the coupling of a spectroscopical technique—micro-Raman spectroscopy—to SEM/EDS is proposed for a sure phase identification of particles, showing EDS spectra with ambiguous phase identification, observed in samples of tissues from patients affected by colorectal cancer and living in an asbestos-polluted area. In these tissues, different particles with EDS spectra that do not allow a sure identification of the phase—in particular calcium-rich particles and titanium oxides—were successively analyzed by micro-Raman spectroscopy. Thanks to this last technique, it was possible to ascribe the mineral phases associated to these particles to “aragonite” (a calcium carbonate polymorph) and to “anatase” (a Ti dioxide polymorph).