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The great potential of graphene can be enhanced thanks to the functionalization of its surface. For this aim, different thicknesses of TiO2 were grown on graphene films by atomic layer deposition (ALD) at 200 °C using H2O and TiCl4 as precursors. The changes in electronic structure of graphene after the deposition of TiO2 and the influence of graphene in TiO2 photocatalytic activity under UV–Visible irradiation were studied. Results indicated the presence of inhomogeneity and intrinsic strain effects within the same sample. Undecorated graphene showed pre-existent strain due to the mismatch between graphene film and the underlying substrate, while non-intentional self-doping is caused by the presence of charged impurities. The deposition of TiO2 films with thickness ≤10 nm led graphene to be p-doped, while strain became the dominant effect increasing film thickness. Oxygen vacancies in the film decreased exponentially by increasing the film thickness leading to a stoichiometric O/Ti atomic ratio of TiO2 above 10 nm thickness. The combination of TiO2 and graphene enhanced the efficiency of electron-hole separation of TiO2 under UV–Visible light, leading to a higher photocatalytic activity tested for methyl red molecule degradation, making TiO2/Graphene hybrid material a promising candidate for the photodegradation of pollutants and water purification.

Bilayer membranes for food packaging applications were prepared by depositing on polylactic acid (PLA) substrates pure cellulose nanofibers' (CNF) coatings and nanocomposite coatings made of CNF with dispersed TiO2 nanoparticles. The gas barrier properties of the bilayer membranes were studied by gas phase permeation technique using CO2, N2, D2, He and ambient air as test gases. Few micrometer thick pure and nanocomposite CNF coatings act as effective gas permeation barriers impeding the CO2, N2 and O2 transport and reducing the He and D2 transport rates by 3 orders of magnitude when compared with the PLA substrate. Gas transport through the coatings obeys the solution-diffusion mechanism: D2 permeation process shows activation energy values of 44 ± 6 kJ/mol and 40 ± 6 kJ/mol in the pure and nanocomposite CNF coating, respectively. The addition of the TiO2 nanoparticles reduces the penetrant diffusivity by increasing the tortuosity of the penetrant migration path but does not affect the gas barrier performances. UV–Vis spectroscopy analyses show that the optical transparency of the PLA support is slightly decreased by the CNF coating: at 600 nm the optical transmittance decreases, in fact, from 95% to 91%. TiO2 nanoparticles strongly absorb UV radiation and slightly reduce the PLA transparency in the optical range due to light scattering effects.

Selective functionalization of the external/internal pore surface of porous silicon is of interest for the numerous potential applications of this material, in particular in pharmacology. With the aim of obtaining porous silicon platforms compatible with the aqueous environment and providing hydrophobic pores to load poorly water soluble molecules, we set-up a three step functionalization procedure consisting in two hydrosilylation reactions separated by the selective etching of the external surface. This procedure was applied both, to porous layers and porous microparticles. The characterization of the functionalized material by conventional techniques such as contact angle and FTIR showed a change of the properties of porous structures in line with the expected surface modifications. However, these techniques do not permit to clearly distinguish between internally and externally grafted functional groups. For this reason, an innovative procedure based on angle-resolved XPS was set-up and applied to differently functionalized pSi layers. By this technique, we obtained indications of prevalent grafting of hydrophilic moieties on the external surface and hydrophobic ones inside the pores.

Mesoporous silicon (pSi) has several interesting features that makes it suitable for various biomedical applications. In particular, the large surface area make it very sensitive to environment changes. Among other approaches, thermal oxidation is an effective way to passivate its surface. Herein, we present experimental and analytical results concerning kinetics of thermal oxidation reaction of pSi. The experiments were conducted on pSi powders produced from silicon wafer by anodization and converted to particles by sonication. Oxidation experiments were carried out at different heating rates. Structure and morphology of the samples have been investigated by XRD and SEM before and after thermal oxidation. The model-free kinetics proposed by Ozawa–Flynn–Wall (OFW) was used to determine the Arrhenius relationship for the pSi thermal oxidation. The obtained apparent activation energy by OFW was confirmed by Starink method. At low temperature, the oxidation of surface dangling bonds obeys the Avrami–Erofeev mechanism. At high temperature, oxidation is followed by classical bulk oxidation according to diffusion mechanism controlled by the diffusion of oxygen through the silicon dioxide layer on the surface of the pSi. The reaction mechanism was checked by the model fitting kinetics, which confirmed the reaction is a kind of sequential two-stage process, Avrami–Erofeev and 3D diffusion. Finally, differential thermal analysis suggests that the second oxidation step is also possibly affected by phase transformation of the silicon dioxide.

α-Tocopherol, the main component of vitamin E, traps highly reactive radicals which otherwise might react with lipids present in plasmatic lipoproteins or in cell membranes. The α-tocopheroxyl radicals generated by this process have also a pro-oxidant action which is contrasted by their reaction with ascorbate or by bimolecular self-reaction (dismutation). The kinetics of this bimolecular self-reaction were explored in solution such as ethanol, and in heterogeneous systems such as deoxycholic acid micelles and in human plasma. According to ESR measurements, the kinetic rate constant (2k(d)) of the bimolecular self-reaction of α-tocopheroxyl radicals in micelles and in human plasma was calculated to be of the order of 10(5) M(-1) s(-1) at 37 °C. This value was obtained considering that the reactive radicals are confined into the micellar pseudophase and is one to two orders of magnitude higher than the value we found in homogeneous phase. The physiological significance of this high value is discussed considering the competition between bimolecular self-reaction and the α-tocopheroxyl radical recycling by ascorbate.

For the first time a variety of foods, characterized by high antioxidant activity, have been tested for the reactivity by which the system of antioxidants present in these foods competes for peroxyl radicals with poly-unsaturated fatty acids. The oxygraphic method we have used, on the basis of a rigorous kinetic model, permits to obtain the reactivity that is the Peroxyl Radical Trapping Efficiency (PRTE), beyond the Peroxyl Radical Trapping Capacity (PRTC) and to assign to each food characteristic values of these parameters, so facilitating their inter-comparison. In the analyzed foods the PRTE/PRTC ratio spans more than one order of magnitude, so reflecting the quality of antioxidants present in foods. According to the PRTE values and on the basis of the serving size the ranking of antioxidant food efficiency in trapping peroxyl radicals was blueberry > red chicory > coffee > pineapple ≈ red wine ≥ orange ≈ dark chocolate ≈ apple ≥ tea > pomegranate.

Porous silicon solid supports with pore diameter 0.5–1 μm, infiltrated with Ag nanostructures for surface enhanced Raman scattering (SERS) were prepared according to two procedures: spontaneous Ag+ reduction on the surface of freshly etched porous silicon immersed in Ag+ aqueous solutions, or anchoring colloidal Ag nanoparticles on the surface previously functionalized by aminosilane. Using Rhodamine 6G (RH6G) as analyte the detection limits were of the order of 20 μM and 20 nM with porous silicon metalized by the first and second procedure, respectively. This large increase of sensitivity notwithstanding a reduced surface density of Rhodamine 6G obtained on porous silicon metalized by the second procedure is discussed in terms of better hot spot efficiency and reduced aspecific binding out of the hot regions obtained depositing the colloids on the aminosilane functionalized surface.

Silver nanocrystallites are obtained through immersion of porous silicon samples in AgNO 3 solutions and a successive thermal annealing. The efficiency of nanostructures as surface enhanced Raman scattering (SERS) substrates is checked on cyanine-based dyes and horseradish peroxidase, evidencing detectable concentrations as low as 10 −7 to 10 −8 M. The substrate efficiency is strictly related to the Ag particle morphology, which could yield to either local surface plasmons (LSP) coupled to individual particles or to inter-particle short-range interaction.

In this paper, we report on a method to evaluate the activity of water soluble and H-atom donor antioxidants as peroxyl radical scavengers in a micelle system reproducing the conditions occurring in the upper small intestine in humans, during digestion and absorption of lipids. This method, which overcomes some of the problems of the total radical trapping antioxidant parameter (TRAP) assays, measures the peroxyl radical trapping capacity (n) and the peroxyl radical trapping efficiency IC 50 − 1 of antioxidants, that is the number “n” of peroxyl radicals trapped by one molecule of the studied antioxidant and the reciprocal of the antioxidant concentration that halves the steady-state concentration of peroxyl radicals, respectively. These two fundamental parameters characterizing the radical chain breaking of many water soluble antioxidants, among which dietary polyphenols, can be obtained with relatively good precision from a single experiment, on the basis of a rigorous treatment of the kinetic data.

The assessment of kidney viability before transplantation (with a view of discarding nonviable organs) remains an obstacle to confidently extending organ harvesting to marginal donors. In the present study phosphorus magnetic resonance spectroscopy was used to monitor metabolic changes in (31)P-containing metabolites in isolated porcine kidneys. After various warm ischemia times, the organs were stored at 0 degrees C. Time-dependent changes in the phosphomonoester/inorganic-phosphate ratio were recorded at 0 degrees C were shown to follow a biexponential decay. The first-order kinetic rate constant of the short-time decay was strongly dependent on the warm ischemia time, a result that was discreted in terms of the underlying biochemistry. The metabolic events responsible for the dramatic decrease in phosphomonoester/inorganic phosphate ratio that occur immediately after organ perfusion and storage, suggest that any procedure to minimize organ damage must occur immediately after harvesting.

The inhibitory effect of anthocyanins has been investigated in the peroxidation of linoleic acid in micelles in the presence and in the absence of (+)-catechin. The peroxidation was initiated by thermal decomposition of 2,2(')-azobis[2-(2-imidazolin-2-yl)propane], and the kinetics of peroxidation were followed by measuring the rate of oxygen consumption and the rate of disappearance of the antioxidant. The analysis of the antioxidant effect of various anthocyanins, alone or in the presence of catechin, demonstrates that catechin, which is relatively inefficient at inhibiting linoleic acid oxidation, regenerates the highly efficient antioxidant malvidin 3-glucoside and, at a lower extent, peonidin 3-glucoside. The malvidin 3-glucoside recycling by catechin strongly increases the antioxidant efficiency of these two antioxidants. This protective mechanism appears specific for malvidin and peonidin 3-glucosides. The high unpaired spin density of the phenolic O atoms in the radicals generated by these anthocyanins, calculated by the semiempirical quantum chemical AM1 method, may explain the observed behavior.

In this paper, we report on the presence of cation binding areas on bovine serum amine oxidase, where metal ions of the groups IA and IIA, such as Na(+), K(+), Cs(+), Mg(2+), and Ca(2+), bind with various affinities. We found a cation-binding area that influences the enzyme activity if occupied, so that the catalytic reaction may be altered by some physiologically relevant cations, such as Ca(2+) and K(+). This binding area appears to be localized inside the enzyme active site, because some of these cations act as competitive inhibitors when highly charged amines, such as spermine and spermidine, are used as substrates. In particular, dissociation constant values (K(d)) of 23 and 27 mM were measured for Cs(+) and Ca(2+), respectively, using, as substrate, spermine, a polyamine of plasma. An additional cation-binding area, where metal ions such as Cs(+) (K(d) congruent with 0.1 mM) and Na(+) (K(d) congruent with 54 mM) bind without affecting the enzyme activity, was found by NMR.

The isoenzyme I of cytosolic Cu,Zn-superoxide dismutase (SOD) from Nicotiana plumbaginifolia (tobacco) leaves has been purified to apparent homogeneity. The relative molecular mass of the native isoenzyme, determined by gel filtration chromatography, is about 33.2 kDa. SDS-polyacrylamide gel electrophoresis shows that the enzyme is composed of two equal subunits of 16.6 kDa The isolectric point, assayed by isoelectric focusing, in the pH range of 3.5–6.5, is 4.3. The enzyme stability was tested at different temperatures, pH, and concentration of inhibitors (KCN and H 2 O 2 ). The catalytic constant ( k cat ) was 1.17 ± 0.14 × 10 9 M −1 s −1 at pH 9.9 and 0.1 M ionic strength. The activation energy of the thermal denaturation process is 263 kJ mol −1 . The electrostatic surface potential of the modeled tobacco Cu,Zn-SOD I was calculated showing that the functional spatial network of charges on the protein surface has been maintained, independently of the amino acid substitution around the active sites.

Dopaminochrome formation is catalyzed by commercially available purified peroxidases (EC 1.11.1.7) such as horseradish, lacto- and myelo-peroxidase using dopamine, hydrogen peroxide or promethazine sulfoxide as substrates. A rat brain fraction (RBF) catalyzes a similar reaction and its catalytic power increases after preincubation with hydrogen peroxide/ascorbic acid. The activity of both the purified enzymes and the RBF preparation is inhibited by carnosine and characterized by excess substrate inhibition. The enzymes recognize different substrates but show the highest affinity for dopamine. The RBF fraction is strongly buffered against oxidation by compounds such as glutathione and by bioreductive enzymes such as DT-diaphorase (EC 1.6.99.2) which can use as a substrate menadione or dopaminochrome. The rat brain dopamine peroxidizing activity appeared to be mostly bound to the synaptosomal fraction. The reaction catalyzed by the purified peroxidases was followed by electron spin resonance spectroscopy and, unlike that catalyzed by RBF, was shown to produce the signal of a transient dopamine-o-semiquinone radical.

This paper reports on a system for sampling atmospheric formaldehyde by dissolution in an aqueous solution followed by the monitoring of the aldehyde using an ion-sensitive field-effect transistor (ISFET) in conjunction with an enzyme specific for this pollutant. Formaldehyde dehydrogenase from Pseudomonas putida was chosen on the basis of its characteristics to be coupled to the ISFET transducer. This enzyme, using oxidized nicotinamide adenine dinucleoside (NAD) as cofactor, catalyses the oxidation of a molecule of formaldehyde with the parallel production of two protons, which are sensed by the ISFET. The working conditions were chosen to obtain a linear response of the sensor up to 200 μM formaldehyde. On the basis of the enrichment obtained by the sampling system, the detection limit of 10 μM formaldehyde in aqueous solution, achieved by the ISFET biosensor, corresponds to an atmospheric concentration of the formaldehyde in the ppb range.

The generation of the superoxide ion in the oxidation of glutathione (GSH) by molecular oxygen has been demonstrated in the absence and in the presence of a catalytic amount of copper ion. An effort has been made to quantify the production of superoxide ion by an NMR method utilising Cu,Zn superoxide dismutase and to correlate it to the reaction mechanism. The oxidation process is first-order with respect to GSH and molecular oxygen, the ratio Δ[GSH]/Δ[O 2 ] being 4.0 ± 0.1. The rate of superoxide ion generation is independent of the GSH concentration and increases linearly with the O 2 concentration. The addition of Cu 2+ increases the rate of GSH oxidation and of O 2 consumption. Also in this case the oxidation process is first-order with respect to GSH and O 2 , while a saturation effect with respect to Cu 2+ is observed. A kinetic scheme involving the formation of a short-living charge-transfer complex between GSH and oxygen is proposed to explain the experimental data.

A systematic study on the disappearance of the electron spin resonance (ESR) signal of nitroxides based on six-or five-membered ring and bearing various charges was carried out in vitro and in vivo. The second-order kinetic rate constants of the reaction of spin probes with ascorbate were measured in vitro at various temperatures in phosphate buffered saline, and the relative activation energies were calculated. Clearance rates of the nitroxide radicals in rat brain homogenates and in blood indicate that the ascorbate contribution to nitroxide removal is about 50-70% in brain and 50-90% in blood. These rates can be easily calculated on the basis of the ascorbate concentration and of the second-order kinetic rate constants measured in phosphate buffered saline. ESR spectra acquired in vivo in rat head and tail, by an L-band resonator, indicated that the nitroxide decay rate is a first-order kinetic process in both domains and that the positively charged nitroxides are not retained in the brain, whereas the anionic and uncharged nitroxides are. Once nitroxides with piperidine ring enter the brain, their decay appears controlled mainly by ascorbate, while the ascorbate has a negligible influence on disappearance in brain of five-membered ring proxyl nitroxides.

A new approach to the preparation of electrochemical biosensors, based on a mixed Sepharose-carbon paste electrode, is described. The bioelectrode is made from carbon paste which is mixed, during preparation, with a mediator and with Sepharose containing an immobilized enzyme. The immobilized enzymes were glucose oxidase, from Aspargillus niger , and amine oxidase from bovine serum and from soybean seedlings. The Sepharose environment, favourable to the enzyme, and the close proximity of the enzyme redox-mediating and sensing sites, permits the required amount of enzyme to be decreased by two orders of magnitude and allows rapid response to the substrate. Response times as short as 15 s have been measured. The microelectrodes are easily fabricated, and the modified carbon paste can be incorporated in the various sensor configurations (micro, flow, etc.) relevant to clinical analysis.

The entry of Ca2+ in rat synaptosomes was followed with a Ca2+-selective electrode. Extracellular ATP is necessary for the entry which is a function of synaptosomal protein, free Ca2+ and glutamate concentrations. Ketamine, glycine and kainate have negligible effect while quisqualate slightly inhibits the uptake of Ca2+ in the presence of glutamate. The added ATP is hydrolyzed by the synaptosomes through an ouabain-insensitive ecto-ATPase affected by the presence of Ca2+, glutamate and, to a slight extent, NMDA.

An improved amino oxidase enzyme electrode has been constructed and applied to the determination of the amount of polyamines present in real samples. The electrode is based on the amperometric detection of H 2 O 2 produced in the enzymatic oxidation of polyamines by amino oxidase. Amino oxidase from soybean seedlings, characterized by an extremely high activity for cadaverine and putrescine, was used. The enzyme was immobilized in an agarose matrix in the presence of glutaraldehyde and bovine serum albumin on the surface of a Pt electrode. Cadaverine, in concentrations between 0.5 and 500 μM, can be quantitatively determined by use of the amino oxidase electrode, the linear calibration range being 0.5–10 μM. The lower detection limit was 0.2 μM and the response time was 15 to 60 s. Putrescine showed similar behaviour. The maximum current response for cadaverine was 5.1 μA/cm 2 , with an apparent Michaelis—Menten constant ( K m ′) of 0.175 mM. The sensor response was stable for more than 32 hours of continuous operation at room temperature and, in the presence of fish or meat homogenates, no change in the signal-to-noise ratio was observed. The long-term stability, pH and temperature response of the biosensor has also been studied.

The events accompanying the inhibitory effect of alpha-tocopherol and/or ascorbate on the peroxidation of soybean L-alpha-phosphatidylcholine liposomes, which are an accepted model of biological membranes, were investigated by electron paramagnetic resonance, optical and polarographic methods. The presence of alpha-tocopherol radical in the concentration range 10(-8)-10(-7) M was detected from its EPR spectrum during the peroxidation of liposomes, catalysed by the Fe3+-triethylenetatramine complex. The alpha-tocopherol radical, generated in the phosphatidylcholine bilayer, is accessible to ascorbic acid, present in the aqueous phase at physiological concentrations. Ascorbic acid regenerates from it the alpha-tocopherol itself. A kinetic rate constant of about 2 X 10(5) M-1 X s-1 was estimated from the reaction as it occurs under the adopted experimental conditions. The scavenging effect of alpha-tocopherol on lipid peroxidation is maintained as long a ascorbic acid is present.

H2O2 was shown to reduce the copper ion of native bovine Cu,Zn superoxide dismutase (superoxide:superoxide oxidoreductase, EC 1.15.1.1) (ECu2+) and to oxidize the reduced enzyme (ECu+). The time-course of these processes was monitored by NMR measurement of the longitudinal relaxation rate of the water protons. A steady-state characterized by the same ratio [ECu2+]/[( EC2+] + [ECu+]) was obtained either by starting from the oxidized or the reduced enzyme. The kinetics of these processes appear to be quite complex, since different reactions between H2O2, or its reaction products, and the enzyme-bound copper control the reaction rate. The solution of the differential equations describing the kinetic processes showed that the oxidation and the reduction of the copper ion by H2O2 are first-order with respect to the copper ion itself only when these processes approach the steady-state. The rate constants of the reduction and oxidation reactions were calculated according to these equations and were found to have comparable values which are in the range 5-80 and 5-45 M-1.min-1, respectively, changing the pH from 5.6 to 7 at 0.21 M ionic strength. This result, together with the dependence of the reaction rates on pH and ionic strength, points to HO2- as the reactive species in both processes, and indicates that the electrostatic control of the access of the peroxide to the active site is the rate-determining step of the two redox reactions.

A short controlled drop-time mercury electrode was utilized to generate the superoxide ion, O 2 − , in aqueous solutions and to study the dismutation processes of this ion. In particular, by the method of the catalytic currents, it was possible to measure the activity of superoxide dismutases, a class of metallo-enzymes present in aerobic cells. The use of this electrode minimizes the contribution of the spontaneous dismutation of the O 2 − to the limiting currents and, as a consequence, it makes enzyme activity measurements possible down to physiological pH values. The advantages with respect to other physicochemical methods are discussed.

The amount of manganese superoxide dismutase (MnSOD) and the activity of copper-zinc superoxide dismutase (CuZnSOD) have been studied in five karyotypically normal human fibroblast strains, using nuclear magnetic resonance (NMR) and polarographic methods. A significant correlation between the two enzyme activities, and a linear increase of MnSOD with the increase of CuZnSOD have been demonstrated. Both enzymes are present in nuclei, mitochondria, lysosome-microsome fraction and cytosol. These findings suggest that the two enzymes dismutate the O-2 cooperatively and that a common genetic control maintains the relative amounts of the two enzymes constant.

The oxidation of ascorbic acid by molecular oxygen was investigated by optical and magnetic resonance methods in the presence and in the absence of copper and iron complexes. The results show that one superoxide ion is generated for each oxidized molecule of ascorbate. The effect of addition of Cu,Zn superoxide dismutase, which at concentrations higher than 10(-7) M halves the oxidation rate, is discussed and a reaction mechanism is proposed. An upper limit for the kinetic rate constant of the reaction between the ascorbate radical and the superoxide ion is reported.