Phd in EARTH SCIENCES

Thesis title: Dissolution in simulated lung fluids and surface reactivity of mineral fibres of environmental and health interest

The toxicity of inhaled mineral fibres results from a complex, multistep process governed by the interplay of their physical, chemical, and structural characteristics, including morphology, chemical composition, surface reactivity, and biodurability. Because of this complexity, the molecular mechanisms underlying fibre toxicity and carcinogenicity are not yet completely understood and remain the subject of ongoing debate. This thesis addresses this tangled issue by using an integrated mineralogical and geochemical multi-analytical approach. The study aims at shedding new light on how crystal-chemical, structural, and morphological features of mineral fibres drive their alteration pathways in solutions simulating biological fluids, and how such surface alterations may, in turn, affect their surface chemical reactivity (Fenton reaction). In addition to asbestos samples (crocidolite, tremolite, and chrysotile), particular attention was devoted to fibrous antigorite, whose pathogenic potential remains poorly constrained, despite its widespread occurrence and environmental relevance. The last part of this work focused on the mineralogical characterization of asbestos bodies embedded in human lung tissue from former asbestos plant workers by synchrotron light source, to gain insight into their formation mechanisms and possible role in asbestos toxicity. The experimental work and the main obtained results are summarized below. The dissolution behaviour and surface modifications of the fibrous minerals were investigated in batch reactors using artificial lysosomal fluid (ALF, pH 4.5), simulating the lysosomal acidic environment of macrophages, and Gamble’s solution (GS, near-neutral pH), representative of the deep lung interstitial fluid. To better assess antigorite biodurability compared to that of asbestos samples, flow-through tests were also performed. Element release into solution was quantified by ICP-OES, while possible changes in fibre morphology, structure, and surface chemistry (including surface Fe speciation) were examined by FE-SEM, XRPD, and XPS, respectively. Experiments performed in ALF and GS revealed that all samples dissolved incongruently in both solutions, with preferential release of cations located at the octahedral M-sites of the amphibole and serpentine structures. However, under the acidic condition amphiboles displayed considerably higher biodurability than serpentines, reflecting their different chemical structural arrangements. Concerning the fibrous serpentines, antigorite and chrysotile exhibited comparable dissolution rates. Nonetheless, antigorite fibres are expected to persist longer in vivo with respect to chrysotile when accounting for their smaller surface area. Notably, the dissolution rate of chrysotile resulted unexpectedly comparable to that of crocidolite at near-neutral pH, highlighting that chrysotile fibres can be efficiently cleared from the lung only if engulfed within the acidic phagolysosomal intracellular sacs. The modulation of fibre surface reactivity (Fenton reactions) following incubation in simulated lung fluids was assessed by monitoring hydroxyl radical generation through EPR spectroscopy. The obtained results demonstrated that, despite surface alteration upon incubation in simulated lung fluids, the ability of the samples in catalysing •OH generation was never suppressed and keeps sustained over time. This is even true for chrysotile fibres, which underwent significant alteration and amorphization at acidic pH, implying that the mineral may exert, similarly to amphibole asbestos, a chronic detrimental action until fibres are fully disintegrated by macrophage attack in vivo. Moreover, the results of this work evidenced the key role of dissolution-driven surface renewal in Fenton reaction potency of UICC crocidolite. Notably, antigorite fibres showed a free radical yield comparable to that of asbestos samples. Therefore, NOA sites containing veins of fibrous antigorite should be regarded as potential sources of toxic fibres during hazard assessment procedures. The mineralogical characterization of asbestos body samples was performed by synchrotron-based X-ray fluorescence and X-ray absorption spectroscopy at the Fe K-edge. The investigation revealed that asbestos bodies are primarily composed of ferrihydrite and goethite in variable proportions. Goethite likely represents the main crystalline product of ferrihydrite transformation under the acidic conditions generated during repeated attempts of fibre phagocytosis by alveolar macrophages. Ferrihydrite, although metastable, can persist for decades within the lung, probably owing to its ability to bind silicates and incorporate exogenous elements from the biological environment that help stabilize its structure. Notably, goethite is among the most cytotoxic and genotoxic Fe oxides, and its presence within asbestos body coatings underscores a potential role in sustaining oxidative stress and contributing to asbestos-related diseases. The results of this work are expected to be of primary importance for future studies aimed at assessing the mechanisms underlying the observed long-term toxicity of mineral fibres.