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HMS materials with Si/Al ratio = 2.5 have been synthesised and characterized. 27 A1 NMR spectra showed minimal occurrence of octahedral phase after template removal. FT-IR and microcalorimetric results showed that, though having the same chemical composition of Y zeolites, such materials exhibit drastically different acidic/basic properties.

Untreated and iron-coated refractory ceramic fibers (RCFs) 1, 3, and 4 were examined for their potential to generate free radicals and to catalyze hydrogen peroxide decomposition in cell-free assays and were compared for cytotoxic and transforming potencies in Syrian hamster embryo (SHE) cell system. Coating with a high quantity of iron increased the capability of RCFs to generate hydroxyl radicals and to catalyze the decomposition of hydrogen peroxide. In the SHE cells, the untreated RCFs had varying ability to induce inhibition of cell proliferation, cytotoxicity (as measured by the colony-forming efficiency, CE) and morphological transformation, in a concentration-dependent manner. According to cytotoxic and transforming potencies, they ranged as follows: RCF3RCF1RCF4. The lethal concentration 50 (LC50; decrease of CE to 50% of controls after 7 d of treatment) expressed per number of RCF3 and RCF1/cm(2) of culture dish was 2.5 x 10(4) and 3.7 x 10(4), respectively, whereas RCF4 was not cytotoxic up to the highest concentration tested (23.7 x 10(4) fibers/cm(2)). At LC50, RCF3 was 1.4-fold more transforming than RCF1, and the weakest, RCF4, induced less than 1% transformation. Iron coating of RCF1 and RCF3 markedly attenuated their cytostatic, cytotoxic, and transforming potencies without a linear concentration-transformation relationship. In contrast, iron coating of RCF4 affected slightly its low transforming potency, although the growth inhibitory effect was reduced. The observed decrease rather than increase in the cytotoxic and transforming potencies of the active samples RCF1 and RCF3 by their coating with large amounts of ferric iron suggests that it is not the quantity or any form of iron on the surface of fibers but the iron, even in trace, in a particular redox and coordinate state that might play a role in the fiber's surface reactivity with regard to the biological material. Surface chemical functions involved in the interaction with the cell could be inactivated by the deposition of a high quantity of Fe(III) on the surface of fibers. Physicochemical studies correlated to biological effects is an approach for understanding the properties of solids related to a given biological response and for elucidating the cellular and molecular mechanisms.

Using various surface modified silica dusts we previously showed that the samples able to generate reactive oxygen species (ROS) induced transformation of Syrian hamster embryo (SHE) cells. The purpose of the present study was to determine in what measure trace iron on the surface of silica particles is involved in the cellular effects, through the formation of ROS. Min-U-Sil 5 quartz (Q) and an amorphous diatomite earth (DE), as well as the same samples pretreated with the Fe chelators ferrozine (fz, 3 mM) and desferrioxamine (dfx, 0.1 mM), were studied for: (i) morphological transformation; and (ii) aberrant mitotic division of SHE cells, which we reported as one of the possible mechanisms of silica-induced cell transformation. In order to determine the nature of the ROS involved in these effects, the antioxidants superoxide dismutase, catalase and mannitol were added to the treated cells. On a per weight basis, DE appeared more cytotoxic and slightly more transforming than Q. The suspensions of Q and DE preincubated in Fe chelators were significantly less transforming and, in particular Q particles, had reduced potency in inducing abnormal mitoses than the untreated samples. Although the amount of Fe mobilized from the silica particles by dfx was greater than that of Fe(II) mobilized by fz, a decrease in transforming potency was largely achieved after pretreatment of particles with fz. In the presence of antioxidants, both the transformation and mitotic aberration frequencies were significantly decreased. The results show that the iron associated with the surface of particles is implicated in the generation of ROS and is partly responsible for the transforming potency of silicas. This study confirms the role of free radical activity in the induction of cell transformation in vitro, suggesting that oxidative stress is one of the mechanisms of silica carcinogenesis.

The mechanism and kinetics of the extraction of iron ions from crocidolite asbestos have been investigated over long periods of time (30 days). Various chelators (endogenous, analytical reagents and iron sequestering drugs) have been employed. The various chelators extracted iron to different extents. Their potency was unrelated to the thermodynamic stability constant but appears linked to a sort of fitting of the molecule at the particle surface. The mechanism of extraction reveals a fast release of surface iron followed by slow ion diffusion, whereby iron flows out of the fibres. Substitution of iron by other ions present in the solution causes structural instability and collapse of the silica framework, confirmed by a parallel release of silicon into the supernatant at the end of the process. The surface shrinks and partly loses its original reactivity, however, new layers are progressively brought to the solid–liquid interface. Conversely, when the original fibres were in contact with iron bound to a weak chelator, such as Fe–NTA, the surface acts as a chelating agent and withdraws iron from the complex. This suggests that in vivo iron cycling may continuously take place at the particle surface. Modifications of the redox state of iron, following incubation in deferoxamine, are evident from cyclic voltammetry. Although deprived of iron, the deferoxamine-treated fibres exhibited a higher amount of redox cycling iron ions than the original ones.

Alterations in the hydrophobic status of particle surfaces have been suggested to modify the toxic properties of ultrafine TiO2. We investigated the acute inflammatory responses and cell damage after intratracheal instillation of surface modified (hydrophilic and hydrophobic) fine (180 nm) and ultrafine (20-30 nm) TiO2 particles 16 h at equivalent mass (1 or 6 mg) and surface doses (100, 500, 600 and 3000 cm2) in rats. Inflammatory response and most enzyme levels were significantly related to the administered surface dose. The hydrophobic surface of the TiO2 particles, achieved by methylation, induced a lower total cell number and influx of neutrophils (PMN) compared to rats instilled with the 1 mg of the untreated, fine or ultrafine TiO2 but the outcomes were not statistically significant. No differences were observed between fine/ultrafine and hydophilic/hydrophobic TiO2 at the high dose (6 mg) or surface dose over 600 cm2. The differences in BAL cellularity at the low dose were reflected in changes in the chemokine MIP-2, but no differences were seen in levels of macrophage cytokines. Considering the large influx of PMN little cell damage was seen when studying enzyme leakage in lavage fluid, although PMNs appeared to be activated as suggested by increased myeloperoxidase (MPO) activity in the lavage fluid. We conclude that the surface area rather than the hydrophobic surface determines the acute, pulmonary inflammation induced by both fine and ultrafine TiO2.

The cytotoxicity of asbestos has been related to its ability to increase the production of reactive oxygen species (ROS), via the iron-catalyzed reduction of oxygen and/or the activation of NADPH oxidase. The pentose phosphate pathway (PPP) is generally activated by the cell exposure to oxidant molecules. Contrary to our expectations, asbestos (crocidolite) fibers caused a dose- and time-dependent inhibition of PPP and decreased its activation by an oxidative stress in human lung epithelial cells A549. In parallel, the intracellular activity of the PPP rate-limiting enzyme, glucose 6-phosphate dehydrogenase (G6PD), was significantly diminished by crocidolite exposure. This inhibition was selective, as the activity of other PPP and glycolysis enzymes was not modified, and was not attributable to a decreased expression of G6PD. On the opposite, the incubation with glass fibers MMVF10 did not modify PPP and G6PD activity. PPP and G6PD inhibition did not correlate with the increased nitric oxide (NO) production elicited by crocidolite in A549 cells. Experiments with the purified enzyme suggest that crocidolite inhibits G6PD by directly interacting with the protein. We propose here a new mechanism of asbestos-evoked oxidative stress, wherein fibers increase the intracellular ROS levels also by inhibiting the main antioxidant pathway of the cell.

Two amphibole asbestos, crocidolite and amosite, have been heated at 400 and 800 °C in order to examine the variations in some surface properties relevant to asbestos toxicity, such as iron mobility, free radical generation, and hydrophilicity. At 400 °C, only the surface is modified, while at 800 °C the crystalline structure partially collapses (X-ray diffraction) with loss in specific surface (Brunauer−Emmett−Teller). Two chelators, ferrozine and desferrioxamine, were employed to evaluate Fe(II) and Fe(III), respectively. More iron was extracted by deferoxamine from crocidolite than from amosite, but when compared per unit surface, the extent of iron removed from the two asbestos was the same. Ferrozine removed more iron from unheated amosite than from crocidolite. Upon heating, because of the oxidation of Fe(II) to Fe(III), less (400 °C) or no (800 °C) iron is removed by ferrozine. The amount of iron removed by deferoxamine is virtually unaffected by heating at 400 °C but dramatically decreases after t...

Refractory ceramic fibres (RCFs) are amorphous fibres made up of various metal oxides (mainly Al2O3 and SiO2), used as high temperature insulation materials. We have examined the surface properties of three RCFs from the TIMA repository, RCF1, RCF3 and RCF4 (RCF1 heated at the expected working temperature), which showed different potentials for cytotoxicity and the induction of morphological transformation in Syrian hamster embryo cells, RCF4 being less active. The degree of surface hydrophilicity, evaluated by adsorption calorimetry, showed that RCF4 is less hydrophilic than RCF1, likely as a consequence of the prior thermal treatment. In order to investigate the effect of thermal treatment on the cellular response to RCFs we have heated (800°C for 24 h) the most active fibre (RCF3) in air until it was converted to a fully hydrophobic material. The cytotoxic (colony forming efficiency) and transforming potencies of heated and unheated RCF3, measured at concentrations between 1 and 16 µg/cm 2 of culture dish for 7 days, were then compared. Both cytotoxicity and potential to transform cells dramatically decreased when fibres were heated, indicating that hydrophobicity blunts the cellular response to RCF3. These results confirm that the state of the surface of RCFs modulates the biological response elicited and that a correlation exists between the degree of surface hydrophilicity and induction of cell damage and transformation. Thermal treatment, by lowering the surface affinity of RCFs for water, inhibits some cell–fibre interactions and decreases the extent of internalization of the fibres.

Microcalorimetric studies on alkylphosphate-butanol and alkylphosphate-butanol-lecithin systems were carried out as a first step to study the role of butanol (used as cosurfactant) in the formation of microemulsions. The enthalpy of solubilization: ΔH, of the same amount of butanol (1·806 mol kg−1) in aqueous sodium monoalkyl-phosphates, with a hydrocarbon chain varying from 6 to 10 carbon atoms, was investigated as a function of alkylphosphate concentration. Measurements were performed in the presence and in the absence of lecithin. The enthalpy values measured upon addition of butanol to the mixtures were negative in all cases. Below the critical micelle concentration (CMC) of the alkylphosphates, the heats of butanol dissolution were found to decrease upon increasing the concentration of the alkylphosphate. This behaviour was related to the dissolution of butanol in water and to the formation of mixed butanol-alkylphosphate and butanol-alkylphosphate-lecithin micelles. Above the CMC essentially constant ΔH values were found. The ΔH measured may be the sum of simultaneous effects—heat of alcohol dissolution and the simple alkylphosphate micelle dissociation or alkylphosphate-lecithin mixed aggregate dissociation and the heat of mixed micelle formation. The ΔH at the plateaus were linearly related to the corresponding CMC values of the alkylphosphates.

Variously modified quartz dusts and one amorphous diatomaceous earth have been compared in their potential to release HO radicals and in their activity in the Syrian hamster embryo (SHE) cell transformation assay. Both original dusts, made up by well-crystallized quartz particles, or by mostly amorphous, variously shaped, silica particles, were active in HO . release, were cytotoxic, and induced morphological transformation in SHE cells. The cristobalite dust, obtained by heating quartz above the phase transition temperature, lost any activity in free radical release, cytotoxicity, and transforming potency. Surface-modified quartz dusts were obtained by a mild etching with HF, by depriving the surface of trace iron with deferoxamine, or by enriching it with iron. The chemical and biological activity decreased in all cases. Both iron-deprived and iron-enriched quartz were nearly inactive. A linear correlation was found between the amount of HO . released by the particles and the transformation frequency. When the SHE cell assay was performed in the presence of mannitol or antioxidant enzymes (superoxide dismutase [SOD] or catalase), the number of transformed cells markedly decreased. This effect was more pronounced for catalase and mannitol than for SOD. HO . release was reduced, but not suppressed, by deferoxamine. All the above results are consistent with the presence of two kinds of surface sites active in HO . release and cell transformation: (1) silicon-based radicals, abundant on freshly ground dusts, which generate the HO . radicals without the superoxide ion as intermediate; and (2) isolated iron centers where the Haber-Weiss cycle takes place, with the superoxide ion as intermediate. The activities of both sites are inhibited by mannitol or catalase, whereas only the last one is inhibited by SOD.

The amphibole minerals amosite and crocidolite were subjected to calcination and to hydrothermal treatment in order to study the effect of these heat treatments on the ability of the minerals to trigger formation of free radicals, which is known to be a main factor causing asbestosis and other asbestos-induced diseases. Free radical activity of the natural and heat treated minerals was studied by using supercoiled DNA (pUC18 plasmid) as a target molecule, and also by means of EPR spectroscopy. It was shown that after calcination of the natural minerals at 1073 K their free radical activity was strongly decreased These results, which may have relevant consequences for asbestos technology, were correlated with concomitant alteration of the structure and surface chemistry of the minerals during calcination.

Free radical generation at the particle/biological fluid interface is one of the chemical processes that contributes to pathogenicity. In order to investigate the role played by iron, fibres of crocidolite asbestos have been modified by thermal treatments to alter their surface iron content. Two radical mechanisms, HO* from H2O2 and cleavage of a C-H bond, which are both active on the original fibres, have been tested on the modified fibres. C-H cleavage is dependent on Fe(II) abundance and location and is suppressed by surface oxidation while HO* release appears independent of the oxidation state of iron. Quartz specimens with different levels of iron impurities have been tested in a similar manner. A commercially available quartz (Min-U-Sil 5) containing trace levels of iron is also active in both tests, but reactivity is not fully suppressed by treatment with desferrioxamine, which should remove/inactivate iron. The radical yield attained is close to the level produced by a pure quartz dust, suggesting the presence of active sites other than iron. Ascorbic acid reacts with both crocidolite and quartz, with subsequent depletion of the level of antioxidant defences when particle deposition occurs in the lung lining layer. Following treatment with ascorbic acid the radical yield increases with quartz, but decreases with asbestos. Selective removal of iron and silicon from the surface may account for the differences in behaviour of the two particulates.

The selective interaction of ascorbic acid with crystalline silica (quartz) has been studied by measuring the ascorbic acid consumption (by means of UV/vis and IR spectroscopy) and the release of silicon when quartz particles or amorphous silica (Aerosil 50) is incubated in ascorbic acid solution. At a physiological ascorbic acid concentration, quartz, and not amorphous silica, reacts, suggesting the formation of a 1:1 silicon-ascorbate complex, while at higher concentrations, the reacting amount of ascorbic acid exceeds the amount of silicon that is released. Silicon tetrahedra bearing free silanols at the quartz surface are selectively attached by ascorbic acid. The particle-derived hydroxyl radical yield in the presence of hydrogen peroxide is increased on ascorbic acid-treated quartz in comparison with the original sample. The results presented herein are relevant because the depletion of ascorbic acid from the lung lining layer and the increased potential in particle-derived free radical generation may both contribute to the oxidative damage following inhalation of crystalline silica.

Chrysotile and crocidolite fibers incubated in normal human plasma (NHP) generated from the C5 component of complement C5a-type fragments that stimulated polymorphonuclear leukocyte (PMN) chemotaxis. Absorption of NHP with antiserum against C5a totally abolished neutrophil chemotactic activity. Asbestos fibers also produced C5a small peptides in the presence of ethylene glycol bis(beta-aminoethyl ether) N,N,N'N'-tetraacetic acid (EGTA) but not ethylene diamine tetraacetic acid (EDTA). Activation of C5 was significantly inhibited when asbestos fibers were pretreated with iron chelators such as sodium dithionite (DTN), deferoxamine (DFX), or ascorbate (AA). Concentration-related inhibition of C5 activation was also observed when asbestos fibers were added concurrently to plasma in the presence of DFX, 1,3-dimethyl-2-thiourea (DMTU), a strong hydroxyl scavenger, or aprotinin (APR), a specific protease inhibitor. Further, chrysotile and crocidolite significantly increased plasma kallikrein activity. Data demonstrate that asbestos-induced C5 activation plays a role in inflammatory reactions characteristic of asbestosis through mechanisms involving iron ions, hydroxyl radicals, and oxidized C5-ike fragments. The ferrous ions present at the asbestos fiber surface trigger this activation and catalyze, via Fenton reaction, the production of hydroxyl radicals, which in turn convert native C5 to an oxidized C5-like form. This product is then cleaved by kallikrein, activated by the same asbestos fibers, yielding an oxidized C5a with the same functional properties as C5a.

The reactive sites at the surface of crocidolite (amphibole asbestos) which may play a role in the toxicity of the fibrous mineral have been investigated by measuring the adsorption of nitric oxide (infrared spectroscopy and adsorption calorimetry) and the release of free radicals in aqueous suspensions. The fibers have been thermally modifiedprogressively heated in vacuo up to 800 °C, calcined in air at 400 °C, and kept under hydrothermal conditions at 200 °Cin order to mimic modifications likely to occur upon weathering. While the crystal structure, as revealed by XRD, was stable up to 600 °C, IR spectroscopy indicated that from 400 to 600 °C OH groups are eliminated and changes in the lattice vibrations occur. Calcination at 400 °C, which oxidizes part of Fe2+ to Fe3+, as evidenced by diffuse reflectance UV−vis spectroscopy, eliminates OH groups and modifies lattice vibrations. The hydrothermal treatment causes oxidation of Fe2+ only to a limited extent. Upon contact with deuterated water, irreversibly...

Department of Health Risk Analysis and Toxicology, Maastricht University, The Netherlands.Oxygen radical generation due to surface radicals, inflammation, and iron release has been suggested as the mechanism of adverse effects of quartz, such as emphysema, fibrosis, and carcinogenic effects. Therefore, we measured iron release, acellular generation of hydroxyl radicals, and oxidative DNA damage and cytotoxicity in rat lung epithelial (RLE) cells by different coal fly ashes (CFA) that contain both quartz and iron. Seven samples of CFA with different particle size and quartz content (up to 14.1%) were tested along with silica (alpha-quartz), ground coal, and coal mine dust (respirable) as positive control particles, and fine TiO(2) (anatase) as a negative control. Five test samples were pulverized fuel ashes (PFA), two samples were coal gasification (SCG) ashes (quartz content

Some humans are often exposed to airborne mineral dusts at the workplace or in daily life. When inhaled, some kinds of mineral dusts can trigger a pathological response of the respiratory system. Silicosis (from silica dust) and asbestosis (from asbestos fibres) are the most commonly known diseases originating from inhaled mineral dusts; other examples are bronchogenic carcinoma and mesothelioma. Detailed knowledge of the chemical (and physical) factors underlying mineral dust toxicity is much needed in order to evaluate the relative risks from exposure to different kinds of materials, both natural and synthetic. These pathogenic factors have been reviewed, with a focus on the surface chemistry of mineral particles and interface phenomena. To facilitate understanding, an outline of the anatomy of the respiratory system and of the etiology of the main diseases involved is also given.

The variability of quartz hazard is related to the characteristics of particulate toxicants. Although these have the same chemical composition, they exist in various forms and surface states, each one eliciting different biological responses. On the basis of data from the literature, surface chemical properties are associated to the subsequent stages reported by Donaldson and Borm (1998) in the mechanistic model proposed for quartz carcinogenicity. Surface radicals and iron-derived reactive oxygen species (ROS) are implicated in oxidative stress, considered to be the key event in the development of fibrosis and lung cancer. Other chemical functionalities related to cytotoxicity, however, modulate the overall pathogenicity by regulating transport and clearance. The chemical features deriving from the intrinsic characteristics of a silica dust--e.g. its origin--as well as those generated by external factors--e.g. contaminants, associated minerals--are discussed in relation to their possible role in the pathogenic mechanism.

The microsomes from dehydroepiandrosterone (DHEA)-supplemented animals are good hydroxyl radical scavengers, as demonstrated through electron spin resonance and deoxyribose degradation. The ability of DHEA-supplemented microsomes to react with superoxide radical was also demonstrated through the inhibition of nitroblue-tetrazolium reduction determined by superoxide radicals produced in a hypoxanthine-xanthine oxidase system. DHEA-enriched microsomes, obtained from acutely DHEA-treated rats, become resistant to iron-dependent lipid peroxidation triggered by H2O2/FeSO4 and ascorbate/FeSO4. The direct addition of DHEA to microsomes from untreated rats failed to prevent iron-dependent lipid peroxidation, even if the microsomes were preincubated with DHEA for up to 15 min, indicating that in vivo transformation is required before antioxidant action can be exerted.

The peculiar characteristics of dust toxicity are discussed in relation to the processes taking place at the particle-biological medium interface. Because of surface reactivity, toxicity of solids is not merely predictable from chemical composition and molecular structure, as with water soluble compounds. With particles having the same bulk composition, micromorphology (the thermal and mechanical history of dust and adsorption from the environment) determines the kind and abundance of active surface sites, thus modulating reactivity toward cells and tissues. The quantitative evaluation of doses is discussed in comparisons of dose-response relationships obtained with different materials. Responses related to the surface of the particle are better compared on a per-unit surface than per-unit weight basis. The role of micromorphology, hydrophilicity, and reactive surface cations in determining the pathogenicity of inhaled particles is described with reference to silica and asbestos toxicity. Heating crystalline silica decreases hydrophilicity, with consequent modifications in membranolytic potential, retention, and transport. Transition metal ions exposed at the surface generate free radicals in aqueous suspensions. Continuous redox cycling of iron, with consequent activation-reactivation of the surface sites releasing free radicals, could account for the long-term pathogenicity caused by the inhalation of iron-containing fibers. In various pathogenicities caused by mixed dusts, the contact between components modifies toxicity. Hard metal lung disease is caused by exposure to mixtures of metals and carbides, typically cobalt (Co) and tungsten carbide (WC), but not to single components. Toxicity stems from reactive oxygen species generation in a mechanism involving both Co metal and WC in mutual contact. A relationship between the extent of water adsorption and biopersistence is proposed for vitreous fibers. Modifications of the surface taking place in vivo are described for ferruginous bodies and for the progressive comminution of chrysotile asbestos fibers.

Carbohydrate moieties of cell glycoconjugates play a pivotal role in molecular recognition phenomena involved in the regulation of most biological systems and the changes observed in cell surface carbohydrates during cell activation or differentiation frequently modulate certain cell functions. Consequently, some aspects of macrophage response to particle exposure might conceivably result from alterations in glycosylation. Therefore, the effect of mineral particles on protein glycosylation was investigated in phorbol myristate acetate (PMA)-differentiated U-937. Jacalin, a lectin specific for O-glycosylated structures, showed a global increase in O-glycosylation in particle-treated cells. In contrast, no significant modifications were observed with concanavalin A, a lectin that recognizes certain N-glycosylated structures. The sialic acid-specific lectins Sambucus nigra agglutinin and Maackia amurensis agglutinin and the galactose-specific lectin Ricinus communis agglutinin revealed a complex pattern of alterations in glycoprotein glycosylation after crystalline silica or manganese dioxide treatments. Expression of sialyl Lewis(x), a glycosylated structure implicated in leukocyte trafficking, could not be detected in control or treated cells. This finding was consistent with the decrease in sialyl Lewis(x) expression observed during PMA-induced differentiation. In conclusion, various treatments used in this study induced quantitative as well as qualitative changes in protein glycosylation. Whether these changes are due to glycosidase release or to an alteration in glycosyltransferase expression remains to be determined. The potential functional implications of these changes are currently under investigation. Images Figure 1. A Figure 1. B Figure 2. A Figure 2. B Figure 3. A Figure 3. B Figure 3. C Figure 4.

The presence of acidic sites on mineral dusts found to be inhibitors of the enzyme human leucocyte elastase (HLE) has been investigated by means of calorimetric and spectroscopic (IR and EPR) techniques. Two acidic phyllosilicates (montmorillonite and kaolinite), which were the most potent inhibitors, and a basic one, muscovite (poor inhibitor) have been examined from the standpoint of their ability to strongly bind ammonia from the gas phase and to adsorb arginine from aqueous solutions. A correlation between inhibiting potency, strong surface acid sites (heat of adsorption of ammonia above 100 kJ mol -* ), and arginine adsorption is envisaged. Chrysotile, a magnesium phyllosilicate, is also a rather poor inhibitor of the enzyme. When the magnesium hydroxide layer is partly leached out, its affinity toward basic molecules and its inactivating potency increase. A model is proposed whereby the enzyme is inactivated when strongly adsorbed on surface acidic sites-dissociated in the aqueous medium-arising from polarized water on small cations (mainly aluminum) and/or particularly acidic surface hydroxyls. This is consistent with the reversibility of the inactivation upon variation of pH and increase in ionic strength.

Hard metal dusts, typically WC/Co, but not pure WC or Co particles, cause the so-called ‘hard metal lung disease’ when inhaled over long periods of time at the workplace. In order to investigate the chemical nature of the dust which originates the disease, the surface behaviour of pure cobalt, pure tungsten carbide, an industrial hard metal dust and a mechanical mixture of cobalt and tungsten carbide have been compared. Electron microscopy reveals an intimate contact between metal and carbide in the mixed dusts. The mixed dust is more active than the single components in the adsorption of water vapour in both adsorbed amount and interaction energy (111 kJ mol -1 for the mixture, 95 kJ mol -1 for pure cobalt and 84 kJ mol -1 for pure WC). Both industrial and mechanical mixtures are more active than pure components in the catalytic decomposition of hydrogen peroxide. Incubation of the mixed dusts in phosphate buffered solutions causes a progressive release of cobalt(ii) ions in solution and the appearance of round smooth aggregates (diameter ca. 300–400 µm) at the expense of smaller particles. The mixed dusts, but not the pure components, promote the homolytic rupture of a carbon–hydrogen bond in aqueous suspension, as revealed by the formation of carboxylate radicals from formate ions. This is evidenced by the use of DMPO as a spin trap, which yields the DMPO–CO 2 - adduct whose EPR spectrum intensity measures the amount of radicals generated. Radicals are only formed in aerated solutions indicating a crucial role of atmospheric oxygen in their generation. The hydroxyl radical, however, does not appear to be implied, for two main reasons: (i) no free oxygen radicals are detected in the absence of formate as target molecule; and (ii) free-radical release is insensitive to the addition of mannitol (an OH scavenger). The formation of the carboxylate radical CO 2 - is an activated process: an induction time of ca. 30 min is required to produce detectable amounts of radicals, while radical generation continues for several hours. Samples withdrawn from the solution, washed, dried and re-employed are still active, as long as some metallic cobalt is present. A model is proposed whereby in the mixture electrons from oxidized cobalt are translocated at the carbide surface where they reduce atmospheric oxygen in a surface-active form which is responsible for the generation of carboxylate radicals from formate ions. The implication of this reaction in health related effects as well as possible hazards from particulates in enviromental pollution is discussed.

The potential for free radical release has been measured by means of the spin trapping technique on three kinds of iron containing particulate: two asbestos fibers (chrysotile and crocidolite); an iron-exchanged zeolite and two iron oxides (magnetite and haematite). DMPO (5,5'-dimethyl-1-pirroline-N-oxide), used as spin trap in aqueous suspensions of the solids, reveals the presence of the hydroxyl and carboxylate radicals giving rise respectively to the two adducts [DMPO-OH] and [DMPO-CO2], each characterized by a well-defined EPR spectrum. Two target molecules have been considered: the formate ion to evidence potential for hydrogen abstraction in any biological compartment and hydrogen peroxide, always present in the phagosome during phagocytosis. The kinetics of decomposition of hydrogen peroxide has also been measured on all solids. Ferrozine and desferrioxamine, specific chelators of Fe(II) and Fe(III) respectively, have been used to remove selectively iron ions. Iron is implicated in free radical release but the amount of iron at the surface is unrelated to the amount of radicals formed. Only few surface ions in a particular redox and coordination state are active. Three different kinds of sites have been evidenced: one acting as H abstracter, the other as a heterogeneous catalyst for hydroxyl radical release, the third one related to catalysis of hydrogen peroxide disproportionation. In both mechanisms of free radical release, the Fe-exchanged zeolite mimics the behaviour of asbestos whereas the two oxides are mostly inert. Conversely magnetite turns out to be an excellent catalyst for hydrogen peroxide disproportionation while haematite is inactive also in this reaction. The results agree with the implication of a radicalic mechanism in the in vitro DNA damage and in the in vivo toxicity of asbestos.

Hard metal alloys (or cemented carbides) are made of a mixture of tungsten carbide particles (WC, more than 80%) cemented in cobalt metal powder (Co, 5-10%). The inhalation of hard metal particles may cause an interstitial pulmonary disease, the mechanism of which involves an interaction between Co and WC particles. Some epidemiological data also suggest that hard metal dust can induce lung cancer in workers. In a macrophage culture model, butylated hydroxytoluene (1 mM) protected from the cytotoxicity of hard metal particles, suggesting a possible involvement of lipid peroxidation in the toxicity of these powders. In a biochemical system, a mixture of Co and WC particles, but not Co or WC alone, stimulated the production of thiobarbituric acid-reactive substances from arachidonic acid. Using a spin trapping system applied to aqueous particulate suspensions and electrochemical techniques, we present experimental evidence that the association of Co and carbide particles represents a specific toxic entity producing large amounts of activated oxygen species. The mechanism of this interaction proceeds through the oxidation of cobalt metal catalyzed at the surface of carbide particles and resulting in the reduction of dissolved oxygen. This physicochemical property of hard metal particles provides a new basis for interpreting their inflammatory action and their possible carcinogenic effect on the lung.

No detectable amounts of iron can be found in simple aqueous suspension of asbestos. In presence of iron chelators, as discovered by Aust and associates (Lund and Aust, 1990; Aust and Lund, 1991; Lund and Aust, 1992; Chao and Aust, 1993 and references therein) considerable amounts of iron are progressively removed from the solid and brought into the solution. It was clearly proved that this mobilized iron was active in DNA damage and lipid peroxidation. The solid may, however, play some role too in asbestos toxicity, acting both as a continuous source of iron ions and as a possible site for deposition of endogenous iron, which under some circumstances may become redox active and consequently toxic. We have therefore set up a thorough investigation on the kind of reactions governing iron release at the solid-liquid interface and on the modifications occurring consequently on the solid. In the present paper we report some results obtained with the three most commonly found asbestos - crocidolite, amosite and chrysotile- and with three different chelators, desferoxamine B, ferrozine and ascorbate. Desferoxamine B, a chelator originated from siderophors, because of its strong affinity for Fe(III), is usually used to block any reaction originated from free iron. Ferrozine, at the opposite, because of its nitrogen donors groups, is a typical analytic reagent for Fe(II). Ascorbate is at the same time a chelator and an endogenous reducing agent, thus its reactivity with the inhaled particle may be relevant to fiber toxicity in vivo.

Once inhaled a particle interacts at various stages, during its path in the respiratory tract, with cells, tissues and biological fluids. Each contact involves a specific functionality at the particle surface, so that the chemical features determining the overall pathogenicity of a given dust are probably more than one. Modulation of one surface property may thus end up in variation of the ultimate toxicity caused even by active surface sites which are unchanged. For this reason investigations on all surface properties of a toxic dust are required.

The hemolytic reaction to a dust is often used as a potential indicator of fibrogenicity of silicon dioxide polymorphs. However, occasionally the hemolytic response may not correlate with the observed fibrotic response in vivo. For example, amorphous silicas are very hemolytic but have little or no fibrogenic activity. In our study, heat treatment was used to alter alpha-cristobalite, a known fibrogenic dust, to a more hydrophobic surface. Comparisons were made between heated and unheated alpha-cristobalite for hemolytic activity in vitro and for lung response in vivo. Heat treatment resulted in decreased hemolytic response, but no change in the fibrotic response occurred in vivo. In addition, the heat treatment resulted in increased initial dust accumulation, reduced short-term clearance, and enhanced long-term clearance in vivo. Increased inflammatory cell recruitment was also observed in lungs of animals exposed to alpha-cristobalite. Thus, whereas heat-induced surface changes in alpha-cristobalite markedly altered the hemolytic activity of the particles, no changes were observed in the fibrotic response.

A preparation of HfO2, derived from the hydrolysis of hafnium isopropylate, has been characterized by XRD, (HR)TEM, MR and adsorption microcalorimetry. The thermal destruction of the amorphous hafnium hydroxide starting phase is complete at ≈700 K, and leads to the crystalline (monoclinic) phase of HfO2. The latter exhibits a particle morphology which, upon thermal treatment, evolves quickly from one of large and loose aggregates of tiny microcrystallites (microcrystalline HfO2) to one made of large single crystallites or of large polyaggregates, in which relatively small ordered microcrystals stack together in a rather disordered fashion (partially sintered HfO2). The evolving morphology of HfO2 is monitored, on a microscopic surface scale, by a varying IR spectrum of surface OH groups and by a varying surface Lewis acidic activity (e.g. towards CO chemisorption), due to coordinatively unsaturated Hf4+ surface centres produced upon vacuum activation. CO uptake, both at ambient temperature and at low temperature (≈78 K), is mainly due to two families of adsorbing sites: sites in structurally and/or coordinatively highly defective configurations, onto which CO adsorbs with an adsorption enthalpy of ≈65 kJ mol−1, and sites located in relatively extended patches of regular crystallographic planes, onto which CO adsorbs with an adsorption enthalpy of ≈50 kJ mol−1. The relative population of the two families of adsorbing sites depends to some extent on the degree of sintering of the material, but it is observed that, unlike other similar systems, the early sintering process causes a rather limited destruction of the crystallographically/coordinatively defective configurations. Both families of adsorbed CO species exhibit a strong dependence of their spectral features on several parameters, among which of primary importance are the degree of surface hydration/dehydration and the surface concentration of charge withdrawing/releasing adsorbed species which, through surface inductive effects, affect the strength of the CO adsorption process.

Taking advantage of the spontaneous polymerisation of eugenol to lignin-like species catalysed by the surface of crocidolite fibres, a procedure is proposed, possibly useful in asbestos removal and disposal, where the polymer avoids the release of airborne fibres and also scavenges ROS (reactive oxygen species).

A safe UV filter may be obtained by inhibiting the photo-generation of free radicals through modification of the surface of TiO(2) nanoparticles with carbon.

Two new asbestiform minerals, balangeroite (Balangero, Italy) and carlosturanite (Val Varaita, Italy), have been investigated from the standpoint of their potential toxicity. Their characteristics have been compared with chrysotile and antigorite with which they are always associated. EPR spectra of the minerals revealed the presence of paramagnetic ions, including Fe3+ and Mn2+, in different crystal configurations. The presence of Fe2+ was shown by the enhancement of the Fe3+ signals after grinding in air. Balangeroite is the richer in both Fe2+ and Fe3+ which are in magnetic interaction with each other, unlike antigorite, in which only isolated and weakly interactive ions are present. Profound modifications of the structures are found upon standing in solutions mimicking the biological environment. This suggests that these minerals may interact in a number of ways in vivo Taking into account both their form and chemical composition they may be regarded as potentially carcinogenic components of fibrous minerals extracted from the Piedmont mines.

Surface dehydroxylation of amorphous and crystalline silicas (quartz dust) has been investigated from the standpoint of the development of hydrophobicity upon thermal treatment. Hydrophobicity occurs when only siloxane bridges and isolated silanols (IR band at ca. 3750 cm–1) are present and is monitored by an enthalpy of adsorption of water lower than the latent heat of liquefaction (44 kJ mol–1). This calorimetric method allows the evaluation of the extent of hydrophilic and hydrophobic patches when both are present at the surface. All silicas develop hydrophobicity upon thermal treatment in vacuo, but quartz is much less easily dehydroxylated than amorphous materials. It is still mainly hydrophilic after outgassing at 1073 K, whereas pyrogenic silicas (Aerosil) become hydrophobic upon outgassing at T < 673 K. Quartz is also characterized by a few very reactive sites (q 90 kJ mol–1), absent on the amorphous specimens. Both these facts might be related to the specific quartz pathogenicity. Rehydroxylation at room temperature of dehydroxylated silicas occurs to a very limited extent. Hydrophilic patches exhibit a marked heterogeneity towards water with an enthalpy of adsorption decreasing from 90 to 44 kJ mol–1. The enthalpy of adsorption approaches 44 kJ mol–1 corresponding to the addition of multilayers of adsorbed water.

The surface chemistry of freshly created surfaces is discussed from the stand point of their potential toxicity. The most reactive sites are surface radicals (dangling bonds) which originate from homolytic cleavage of covalent bonds and transition metal ions in low oxidation states. Several types of mineral fibres will be discussed: SiC whiskers, asbestos, nemalite, glass and rockwool fibres.

Two surface dinuclear oxygen species simultaneously form when quartz is ground in a solution of hydrogen peroxide. They are the superoxide O − 2 radical ( g zz =2.044, g yy =2.010, g xx =2.002) and the peroxy SiOO· radical ( g zz = 2.064, g yy = 2.007, g xx = 2.002). They are formed by contact of the freshly created surface, bearing reactive centres originated by cleavage of the SiOSi bonds, with H 2 O 2 . In parallel, OH· radicals are detected in the solution. A scheme involving the catalytic decomposition of hydrogen peroxide is proposed accounting for the radical species observed at the surface and in solution.

Stable radicals detectable by electron paramagnetic resonance (EPR) may be use in the investigation of early events in cell-particle toxicity. Piperidine-N-oxyl derivatives (nitroxides), covalently linked to the surface of a high surface area silica (used as model solid for the technique), served as probes in the investigation of the effects of incubation of silica particles with mesothelial cells. A mesoporous silica (MCM-41), prepared by precipitation from a micellar solution, was the most appropriate silica-based particle for this purpose, as its channels allow direct contact with small molecules but not with macromolecules. The cytotoxicity of this amorphous silica is very low, allowing relatively high particle loading in the cell cultures. Both the high surface area of the sample and the large amount of inorganic material extracted from the cell culture provide enough material to run reasonably intense EPR spectra. Computer-aided analysis of the EPR spectra of silica-bound nitroxides provided information on the sensitivity of the labeled silica monitoring different environments, e.g., to follow the path of particles in a mammalian cell culture. Upon contact of the particles with mesothelial cells, the mean distance among the labels at the silica surface decreased as a consequence of the release of oxidizing and/or radical moieties from the cells. Images Figure 5. a Figure 5. b

Stishovite, the SiO2 polymorph obtained at extremely high pressure with octahedrally coordinated Si atoms, has been characterized by means of techniques usually employed in surface chemistry. Its surface properties have been investigated in order to explain why stishovite is the only non-pathogenic SiO2 crystalline polymorph. X-Ray diffraction and HRTEM studies indicate that, instead of promoting a phase transition towards stable polymorphs, heating progressively destroys the crystalline structure, and the solid undergoes a nearly complete amorphisation. Photoacoustic and FTIR spectroscopies show that, on heating from room temperature to the treatments at higher temperatures (up to ca. 1073 K): (i) the unusual octahedral coordination of Si atoms in the stishovite structure is gradually lost, and new absorptions appear due to different νSi—O modes; (ii) the surface OH groups, almost all of which are involved in strong H-bonding interactions in the original material, are different from those in most silicas (no free silanols), and become more and more similar to those of ordinary SiO2 polymorphs. Both FTIR spectra and microcalorimetric data, relative to the adsorption of H2O on stishovite thermally activated in the 300–1073 K range, monitor the presence of a strong surface field: in particular, the type and strength of the surface sites are qualitatively different, and quantitatively stronger, than those usually observed for most other SiO2 polymorphs.

Infrared, volumetric and calorimetric data for TiO2, ZrO2 and HfO2, variously pretreated, are reported and discussed. Two types, at least, of M4+ sites are potentially present on all oxides, differing by the degree of coordinative unsateration. Surface impurities may alter or suppress them. Besides this structural heterogeneity, transmission of electronic effects through the solid induces heterogeneity during adsorption (shift of CO stretching frequency with coverage). The two phenomena are strictly related: the same relationship is proposed for the changes in absorption coefficient with peak position. This allows a new way of decomposing overall isotherms into individual contributions.

Reactive sites on α-cristobalite and α-silicon carbide outgassed over a wide range of temperature (423–1573 K) have been studied by adsorption microcalorimetry. By successive adsorption–desorption cycles the fraction of each adsorbate reversibly adsorbed at room temperature and the effect of preadsorption of one adsorptive on the adsorption of the other have been evaluated. Both molecules are dissociated on strained bridges and adsorbed via H-bonds on surface hydroxyls, but they do not react strictly with the same sites. H-bonding of H2O takes place on two silanols located at a defined distance, with an enthalpy in the range 50–100 kJ mol–1, and on isolated silanols, with an enthalpy lower than the latent enthalpy of liquefaction (44 kJ mol–1). The two processes discriminate hydrophilic and hydrophobic patches at the surface. Ammonia interacts with single silanols, either isolated or terminal in a cluster: in the latter case electronic effects due to adsorption involve all SiOHs in mutual interaction. Strained siloxane bridges dissociate both molecules with formation either of two new silanols or of one silanol and one silylammine group. This latter surface functionality interacts with H2O, although with a lower enthalpy than SiOH, but does not interact with NH3. Silicon carbide exhibits similar behaviour to silicas, when outgassed up to 773 K, but after heating at 1073 K develops a peculiar reactivity, whereby large amounts of both H2O and NH3 are dissociated, in contrast to silica whose reactivity is progressively depressed upon thermal treatments. The above data are discussed in the context of systematic surface characterization of covalent solids, aiming to relate the surface properties of the particles to their lung toxicity when inhaled.