Wine putrescine abatement by bentonites: From ideal case to practice.

Herein, we demonstrated that bentonites can be incisively used to reduce wine BAs content, especially putrescine molecules. Pioneering kinetic and thermodynamic studies of putrescine adsorption onto two commercially available bentonites (optimal concentration of 0.40 g dm-3) were performed resulting in ca. 60% removal by physisorption mechanism. Both bentonites showed also promising results in more complex systems, resulting in a lower putrescine adsorption due to the competition with other molecules (as proteins, polyphenols), typically present in wines. Nonetheless, we managed to reduce the putrescine content below 10 ppm both in red and white wines.

A comprehensive study on the effect of bentonite fining on wine charged model molecules.

In bottled wines, haze and turbidity are phenomena to be avoided. Since bentonite fining is a common process to clarify wines removing heat unstable proteins, a theoretical study on the adsorption of three Charged Model Molecules (CMMs, egg albumin, polyphenols and riboflavin) was carried out to deep comprehend this chemical phenomenon. Four bentonites were adopted and finely characterized together with the potential release of Na+ and Ca2+ cations, revealing suitable for RT albumin removal within 120 min. Better results in terms of adsorbed quantity were achieved by adopting 12%v/v EtOH/H2O solvent and by swelling bentonites for 24 h before use. With the most performing sample (Na/Ca_0.27), a comprehensive study on simultaneous adsorption of the three CMMs was performed, resulting in polyphenols adsorption increase due to their interactions with albumin. Notwithstanding the majority of albumin and riboflavin was successfully removed, ca. 40-50% of tested polyphenols was preserved.

3D Mass Spectrometry Imaging Reveals a Very Heterogeneous Drug Distribution in Tumors.

Mass Spectrometry Imaging (MSI) is a widespread technique used to qualitatively describe in two dimensions the distribution of endogenous or exogenous compounds within tissue sections. Absolute quantification of drugs using MSI is a recent challenge that just in the last years has started to be addressed. Starting from a two dimensional MSI protocol, we developed a three-dimensional pipeline to study drug penetration in tumors and to develop a new drug quantification method by MALDI MSI. Paclitaxel distribution and concentration in different tumors were measured in a 3D model of Malignant Pleural Mesothelioma (MPM), which is known to be a very heterogeneous neoplasm, highly resistant to different drugs. The 3D computational reconstruction allows an accurate description of tumor PTX penetration, adding information about the heterogeneity of tumor drug distribution due to the complex microenvironment. The use of an internal standard, homogenously sprayed on tissue slices, ensures quantitative results that are similar to those obtained using HPLC. The 3D model gives important information about the drug concentration in different tumor sub-volumes and shows that the great part of each tumor is not reached by the drug, suggesting the concept of pseudo-resistance as a further explanation for ineffective therapies and tumors relapse.

Photocatalytic degradation of toluene in the gas phase: relationship between surface species and catalyst features.

Photocatalytic activity of both commercial and homemade samples was tested for the degradation of toluene in the gas phase by using two different irradiation sources, UV and solar. The role played by humidity in affecting the final toluene degradation was discussed. Catalyst deactivation (due to the high toluene level, 1000 ppm) and subsequent regeneration, by washing with water, were analyzed. Highest degradations and corresponding kinetic constants were achieved in the case of the anatase/brookite composite samples, while the commercial ones (including P25 Degussa) showed lower efficiency. Various adsorbed aromatic species (benzoic acid, the major surface product hydroquinone, benzylic alcohol, benzaldehyde, and cresols) obtained by washing the exhaust catalysts were analyzed by HPLC. Parallel results were achieved by Fourier transform infrared attenuated total reflectance (FTIR-ATR) spectroscopy. The presence of different photodegradation surface species for the various photocatalysts suggests the occurrence of different reaction pathways, depending on the fine physicochemical features of the specific TiO2 adopted in the reaction.

Influence of MPP+ on the state of tubulin polymerisation in NGF-differentiated PC12 cells.

Cytoskeletal proteins have been reported as constituents of cytoplasmic inclusions typical of degenerated neurones in Parkinson's disease and, in addition, the involvement of cytoskeleton in the mechanism of action of the parkinsonism-producing neurotoxin MPP+ is emerging. Here we investigate the influence of MPP+ on the dynamic behaviour of microtubules. Neurone-like cells derived from a rat pheochromocytoma cell line (PC12) and differentiated with nerve growth factor are used as a model system. We found that sublethal doses of the neurotoxin markedly affect the state of tubulin polymerisation: polymerised tubulins significantly decreased, whereas an increase of unpolymerised alpha-tubulin was observed. Since the concentration of unassembled tubulin directly regulates tubulin synthesis by a feedback mechanism, we studied alpha- and beta-tubulin synthesis by metabolic labelling of PC12 cells with [35S] methionine and following immunoprecipitations. The results showed the significant decrease of labelling in both the microtubule subunits in cells exposed to the neurotoxin. We suggest that the MPP+-induced imbalance of tubulin polymerisation and synthesis represents a novel early step in the mechanism of action of the neurotoxin.

Apoptosis in human lung epithelial cells: triggering by paraquat and modulation by antioxidants.

Recent results have shown that apoptosis is an important feature of the normal and injured lung epithelium, but little conclusive evidence is available about the exact intracellular mechanisms involved. In this work, we studied apoptotic cell death in the established human lung epithelial cell line, A549, by evaluating the ability of the pulmonary toxin, paraquat (1,1'-dimethyl-4, 4'-bipyridylium dichloride), to act as a trigger, and assessing the ability of ascorbic acid and N-acetyl-cysteine (NAC) to modulate the process. The analysis of nuclear and cellular morphology along with TUNEL staining showed that paraquat is an inducer of apoptosis. A549 cells incubated with sublethal doses of paraquat for up to 24 h showed no apoptotic features but, their following incubation in paraquat-free medium resulted in a time-dependent appearance of apoptosis. The antioxidants, ascorbic acid and NAC, proved effective in reducing paraquat-induced apoptosis, and therefore were seen as protective agents. Finally, we propose an experimental model for investigating some of the key steps in the apoptotic programme in alveolar cells.

Dynamic changes in the state of actin polymerization in human alveolar cells exposed to the oxidant agent paraquat.

To investigate the role of dynamic changes in actin cytoskeleton in cellular response to oxidative stress, we have analyzed the state of actin polymerization and synthesis in human alveolar cells exposed to paraquat, an oxidant agent. Cellular content of monomeric actin was assayed by DNase I inhibition. It decreased significantly in treated cells and depended on paraquat concentration. Paraquat treatment of cells caused an increase of the filamentous pool of actin and a parallel decrease of the monomeric one. Such shift was shown to be irreversible. SDS-PAGE of cytoskeletal fractions was performed under reducing and non-reducing conditions. No cross-linking of actin monomers to form large aggregates appeared to be related to the observed paraquat-induced increase of the filamentous actin pool. Morphological analyses by indirect immunofluorescence and ultrastructural examination confirmed the presence of microfilaments in treated cells. Conventional bundles of filaments were not observed, but numerous single filaments appeared dispersed within the cytoplasm. Pulse-chase experiments showed a strong increase of de novo synthesis of actin in treated cells, whereas actin degradation rate remained unaffected. In conclusion, the different approaches lead to a concordant picture of cellular response to oxidant stress at the level of the actin filament system. Actin pools are modified: the overall number of filaments increases, whereas the monomeric species decreases. As a result of the shift of actin from the monomeric pool to the filamentous one, the de novo synthesis of actin is increased.

Involvement of tubulin in MPP+ neurotoxicity on NGF-differentiated PC12 cells.

In vivo, the neurotoxin MPTP is oxidated to MPP+, which is toxic to dopaminergic neurons. In this paper, we have used MPP+ as a tool to evoke neurotoxicity in the PC12 cell line and investigate the intracellular events that are involved. A cytotoxicity test, performed on undifferentiated and NGF-differentiated PC12 cells, showed that MPP+ is much more toxic on differentiated cells and indicated the suitable range of concentrations for studying the starting events evoked by the neurotoxin. By indirect immunofluorescence we have shown that the localisation of alpha- and beta -tubulin in NGF-differentiated cells was modified by a 24 h treatment with 15 mumol/l MPP+. A biochemical analysis was performed on cell extracts and the results showed that MPP+ treatment induced an increase in alpha -tubulin levels and a decrease in beta -tubulin levels. These results suggest the involvement of the two microtubule proteins in MPP+ neurotoxicity on NGF-differentiated PC12 cells.

Paraquat induces irreversible actin cytoskeleton disruption in cultured human lung cells.

The herbicide paraquat (PQ) induces the selective necrosis of type I and type II alveolar pneumocytes. We investigated the effect of PQ on human lung A549 cells to determine the possible role of cytoskeleton in lung cytotoxicity. At 80 mumol/L PQ, a concentration that did not affect cell viability, the organization of actin cytoskeleton network depended on incubation time with the herbicide. Microfilaments appeared less numerous in 30% of the cells treated for 1 h. After 24 h, all the treated cells displayed only short filaments in the periphery. The effect of PQ on actin cytoskeleton was irreversible. Moreover, no modification of microtubule network was observed in PQ-treated cells. Next, we studied the effect of PQ on Chang Liver, an epithelial cell line from human liver. These cells appeared less sensitive to the herbicide than A549, and no cytoskeletal alteration was observed. To verify whether actin filament modifications in A549 cells were related to intracellular alterations of ATP concentrations, nucleotide levels during incubation with PQ were determined. The intracellular levels of ATP were not different in control and treated cells. Our results indicate that PQ induces specifically an irreversible actin filament disorganization on A549 cells and that the observed effect is independent of intracellular concentration of ATP.