The Antioxidative Effect of Fullerenes during the Peroxidation of Methyl Linoleate in Toluene

The antioxidative effect of fullerenes C₆₀ and C₇₀ was examined by measuring the inhibition of methyl linoleate (MeL) peroxidation in toluene initiated by 2,2′-azobis(2,4-dimethylvaleronitrile) (AMVN). The fullerenes retarded the formation of MeL hydroperoxides and lowered the rate of propagation. The reaction rates of fullerenes with AMVN-derived peroxyl radicals were much higher than that of MeL. These results indicate that fullerenes can act as retarders of lipid peroxidation, though their activity is low compared with that of α-tocopherol.     

Reaction products of [60]fullerene during the autoxidation of methyl linoleate in bulk phase

The antioxidative action of fullerenes has received much attention, but their reaction mechanism toward lipid-derived peroxyl radicals has not been well elucidated. In this study, the reaction products of [60]fullerene (C(60)) during the autoxidation of methyl linoleate (MeL) were isolated and their structures were characterized. MeL containing 0.1mol% C(60) was autoxidized at 60°C in bulk phase and two reaction products of C(60), 1 and 2, were obtained. The structure of 1 was the addition products of C(60) with 9-peroxyl-10-alkyl radicals of methyl (11E)-13-hydroperoxy-11-octadecaenoate (1a and 1b) and with 12-alkyl-13-peroxyl radicals of methyl (10E)-9-hydroperoxy-10-octadecaenoate (1c and 1d). 2 was a mixture of the addition products of C(60) with 9,11-dialkyl radicals of methyl 9,12-octadecadienoate (2a) and with 11,13-dialkyl radicals of methyl 9,12-octadecadienoate (2b). When MeL containing 0.1mol% C(60) was autoxidized at 60°C under air-sufficient and air-insufficient conditions, C(60) could suppress the formation of MeL hydroperoxides in both conditions. The reaction product of C(60) first formed was 2 even under air-sufficient conditions, and then 1 was accumulated. The results indicate that the primary antioxidative action of C(60) would be trapping of chain-initiating carbon-centered radicals of unsaturated lipid to form 2.     

Oxidative properties of photoexcited fullerenes C60 and C60-containing composites in suspension of thymocytes and Ehrlich ascites carcinoma cells

Biological effects of photoexcited in UV/VIS region water-soluble fullerenes C60 and synthesized C60-containing composites after its addition to cell incubation medium are studied. It is shown by EPR method, that in the presence of photoexcited C60 (10(-5) M) reactive oxygen species (ROS) are generated both in water solution and cell suspensions. ROS generation is accelerated if C60 is included into aminopropylaerosyl or antracenaliminopropylaerosyl. Photodynamic damage of Erlich ascites carcinoma cells by photoexcited C60-composites is confirmed by a decrease of cell viability, enhanced content of Schiff base fluorescent products of malonaldehyde as indicator of lipid peroxidation and the decrease in respiratory activity, estimated by MTT-test. No effects of photoexcited C60-composites were detected in thymocytes suspension. The data obtained testity to perspective utility of C60-composites for optimization of photodynamic therapy methods.     

Oxidative stress generated damage to DNA by gastrointestinal exposure to insoluble particles

There is growing concern that gastrointestinal exposure to particles is associated with increased risk of toxicity to internal organs and carcinogenicity. The mechanism of action is related to particle-induced oxidative stress and oxidation of DNA. Observations from animal models indicate that gastrointestinal exposure to single-walled carbon nanotubes (SWCNT), fullerenes C60, carbon black, titanium dioxide and diesel exhaust particles generates oxidized DNA base lesions in organs such as the bone marrow, liver and lung. Oral exposure to nanosized carbon black has also been associated with increased level of lipid peroxidation derived exocyclic DNA adducts in the liver, suggesting multiple pathways of oxidative stress for particle-generated damage to DNA. At equal dose, diesel exhaust particles (SRM2975) generated larger levels of 8-oxo-7,8-dihydro-2'-deoxyguanosine in rat liver than carbon black (Printex 90) did, whereas exposure to fullerenes C60 and SWCNT was the least potent. This ranking of samples was also observed for oxidatively damaged DNA in cultured cells. The extent of translocation from the gut is largely unresolved. However, there is evidence indicating that gastrointestinal exposure to particulate matter is associated with oxidative damage to DNA and this might be associated with increased risk of cancer.     

Ligand effects in cis-Ru(N-N)2L2 species where N-N is a substituted 2,2′-bisoxazoline and L is monodentate

Starting from previously reported cis-Ru(MeL)₂Cl₂, where MeL is 4,4,4′,4′-tetramethyl-2,2′-bisoxazoline, cis-Ru(MeL)₂Br₂ (1), cis-Ru(MeL)₂I₂ (2), cis-Ru(MeL)₂(NCS)₂ · H₂O (3), cis-Ru(MeL)₂(N₃)₂ (4) and cis-[Ru(MeL)₂(MeCN)₂](PF₆)₂ · (CH₃)₂CO (5) are synthesised. The X-ray crystal structures of complexes 1, 2, 3 and 5 have been determined. All the five new complexes have been characterized by FTIR, ESIMS and ¹H NMR. In cyclic voltammetry in acetonitrile at a glassy carbon electrode, the complexes display a quasireversible Ru(II/III) couple in the range 0.32-1.71 V versus NHE. The Ru(II/III) potentials yield a satisfactorily linear correlation with Chatt’s ligand constants P_L for the monodantate ligands. From the intercept and by comparing the known situation in Ru(2,2′-bipyridine)₂L₂, it is concluded that MeL, a non-aromatic diimine, is significantly more π-acidic than 2,2′-bipyridine.     

Antiamyloid properties of fullerene C60 derivatives

A comparative estimation of the ability of complexes of fullerene C60 with polyvinylpyrrolidone and fullerene C60 derivatives (the sodium salt of the polycarboxylic derivative of fullerene C60, sodium fullerenolate), has been carried out. The fullerenes destroyed amyloid fibrils of the Abeta(1-42) peptide of the brain and the muscle X-protein. A study of the effect of fullerenes on muscle actin showed that complexes of fullerene C60 with polyvinylpyrrolidone and sodium fullerenolate did not prevent the filament formation of actin, nor did they destroy its filaments in vitro. Conversely, sodium salt of the polycarboxylic derivative of fullerene C60 destroyed actin filaments and prevented their formation. It was concluded that sodium fullerenolate and complexes of fullerene C60 with polyvinylpyrrolidone are the most effective antiamyloid compounds among the fullerenes examined.     

Novel C5a Agonist-Based Dendritic Cell Vaccine in a Murine Model of Melanoma

A novel method to improve targeting and presentation of poorly immunogenic tumor-related antigens was investigated. This was performed with a molecular adjuvant constructed by covalently linking a response selective peptide agonist of C5a (YSFKDMP(MeL)aR) to known melanoma tumor-related antigens. C57Bl/6J mice were injected subcutaneously with bone marrow derived dendritic cells (DCs) pulsed with a melanoma epitope (TRP2-P2/Agonist), melanoma epitope tyrosinase (TYR/Agonist), a nonfunctional reverse conformation C5a agonist bound to TYR(reverse peptide) or DMSO-PBS vehicle. Mice were injected with the pulsed DCs and cytokines IL-2 and GM-CSF three times prior to subcutaneous challenge with B16-F10 melanoma cells. All groups subsequently received DC vaccine boosters twice per week. Tumor growth was reduced and survival enhanced in mice immunized with the combination of TRP2-P2/Agonist and TYR/Agonist compared to mice receiving reverse peptide or vehicle.     

Generation of long-lived radical ions through enhanced photoinduced electron transfer processes between [60]fullerene and phenothiazine derivatives.

Photoinduced electron transfer processes between fullerenes (C60) and four phenothiazine derivatives (PTZs) in the absence and presence of hexylviologen dication (HV2+) have been studied by the transient absorption method in the visible and near-IR regions. Electron-transfer takes place from PTZs to the triplet states of fullerenes (3C60*) giving the radical anion of fullerenes (C60.-) and the radical cations of PTZs (PTZ.+). The rate constants and efficiencies of electron transfer are quite high, because of the high electron-donor abilities of PTZs as elucidated by their low oxidation potentials. On addition of HV2+ to the C60 and PTZ systems, the electron-mediating process occurs from C60.- to HV2+, yielding the viologen radical cation (HV.+). In the presence of a sacrificial donor, HV.+ persisted for a long time.     

Direct preparation of radioactive fullerenes as a tracer for applications

The C₆₀ and C₇₀ fullerenes were irradiated by high-energy γ-rays and charged particles. The irradiated samples were dissolved in CS₂ and/or toluene and filtered to remove insoluble by-products. Finally, radioactive fullerenes and products labeled with¹¹C or¹³N were isolated and detected in the liquid phase by radiochromatography. It was found that (1) not only¹¹C or¹³N radioactive monomer fullerenes but also their dimers (trimers and, possibly, tetramers) were produced by recoil implantation process following nuclear reaction and (2) the radioactive fullerene labeled with¹¹C yields has led to high yields.     

Chronic alcoholization-induced damage to astroglia and intensification of lipid peroxidation in the rat brain: Protector effect of hydrated form of fullerene C<Subscript>60</Subscript>

We studied the intensity of lipid peroxidation (LP) and the amount of a marker of astrocytes (glial fibrillary acidic protein, GFAP) in tissues of the rat brain under conditions of long-lasting consumption (12 weeks) of ethyl alcohol, as well as the protective effects of peroral administration of hydrated forms of fullerene ?₆₀ (?₆₀HyFn, FWS, fullerene water solutions). Consumption of ethanol resulted in a rise in the amount of molecular markers of oxidative stress (thiobarbiturate-active compounds) in the cerebral tissues. The level of the filamentous GFAP form in the hippocampus and cerebral cortex of alcoholized animals decreased significantly, which can be a result of death of the population of GFAP-imunnoreactive astrocytes in the brain. In the brain of rats after systematic consumption of both ethanol and an aqueous solution of hydrated fullerenes ?₆₀, the amounts of products of lipid peroxidation and of the astroglial marker did not differ significantly from the respective indices in the control animals. Our data demonstrate the efficiency of hydrated fullerenes as pathogenetic therapeutic remedies for elimination of the negative effects of ethyl alcohol on the CNS.     

Protein interaction with hydrated C60 fullerene in aqueous solutions

Physicochemical effects of hydrated C(60) fullerenes (HyFn) on serum albumin molecules were studied using ESR spin labeling and differential scanning microcalorimetry. Molecular-colloidal solution of hydrated C(60) fullerenes and their small spherical fractal clusters in water (C(60)FWS), was shown to stabilize protein hydration, and decrease specific surface energy in water-protein matrix in salt solutions. The mechanism of HyFn interaction with protein is discussed in terms of HyFn induced formation of protein clusters and phase transition of hydration water.     

C60 binds to and deforms nucleotides

Atomistic molecular dynamics simulations are performed for up to 20 ns to monitor the formation and the stability of complexes composed of single- or double-strand DNA molecules and C60 in aqueous solution. Despite the hydrophobic nature of C60, our results show that fullerenes strongly bind to nucleotides. The binding energies are in the range -27 to -42 kcal/mol; by contrast, the binding energy of two fullerenes in aqueous solution is only -7.5 kcal/mol. We observe the displacement of water molecules from the region between the nucleotides and the fullerenes and we attribute the large favorable interaction energies to hydrophobic interactions. The features of the DNA-C60 complexes depend on the nature of the nucleotides: C60 binds to double-strand DNA, either at the hydrophobic ends or at the minor groove of the nucleotide. C60 binds to single-strand DNA and deforms the nucleotides significantly. Unexpectedly, when the double-strand DNA is in the A-form, fullerenes penetrate into the double helix from the end, form stable hybrids, and frustrate the hydrogen bonds between end-group basepairs in the nucleotide. When the DNA molecule is damaged (specifically, a gap was created by removing a piece of the nucleotide from one helix), fullerenes can stably occupy the damaged site. We speculate that this strong association may negatively impact the self-repairing process of the double-strand DNA. Our results clearly indicate that the association between C60 and DNA is stronger and more favorable than that between two C60 molecules in water. Therefore, our simulation results suggest that C60 molecules have potentially negative impact on the structure, stability, and biological functions of DNA molecules.     

Using C60 fullerenes for photodynamic inactivation of mosquito iridescent viruses

This article describes the photodynamic inactivation of mosquito iridescent virus (MIV) Aedes flavescens in the presence of water-soluble C(60) fullerenes. It has been observed that the photodynamic inactivation of MIV for about 1 h reduces the infectious titre of the virus in large wax-moth larvae Galleria mellonella to 4.5 lg ID(50)/mL. The influence of the C(60) concentration on its anti-viral activity was tested in the concentration range from 1 to 0.001 mg/mL. It has been found that C(60) is able to inactivate the iridovirus even in low concentrations. Consequently, the findings of this work suggest that photoexcited C(60) fullerenes can be successfully used for the inactivation of iridoviruses in biological systems.     

Effect of lipid physical state on the rate of peroxidation of liposomes.

The effect of cholesterol on the rate of peroxidation of arachidonic acid and 1-palmitoyl-2-arachidonoyl phosphatidylcholine (PAPC) in dimyristoylphosphatidylcholine (DMPC) liposomes was examined above and below the phase transition temperature (Tm) of the lipid. The rate of peroxidation of arachidonic acid was more rapid below the phase transition temperature of the host lipid. At a temperature below the Tm (4 degrees C), increasing concentrations of cholesterol reduced the rate of peroxidation of arachidonic acid as judged by the production of thiobarbituric acid reactive substances. Above Tm (37 degrees C), cholesterol increased the rate of peroxidation of the fatty acid. Similarly, PAPC was peroxidized more rapidly at 4 degrees C than at 37 degrees C. However, cholesterol had little effect on the rate of peroxidation of PAPC at 4 degrees C. The rate of peroxidation of arachidonic acid was related to the lipid bilayer fluidity as judged by fluorescence anisotropy measurements of diphenylhexatriene. The rate of peroxidation increased slowly with increasing rigidity of the probe environment when the bilayer was relatively fluid and more rapidly as the environment became more rigid. The increase in the rate of peroxidation of arachidonic acid in the less fluid host lipid was unrelated to differences in iron binding or to transfer of arachidonic acid to the aqueous phase. Decreasing the concentration of arachidonic acid in DMPC to less than 2 mol% dramatically decreased the rate of peroxidation at 4 degrees C, suggesting that formation of clusters of fatty acids at 4 degrees C is required for rapid peroxidation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Fullerene nanomaterials inhibit the allergic response

Fullerenes are a class of novel carbon allotropes that may have practical applications in biotechnology and medicine. Human mast cells (MC) and peripheral blood basophils are critical cells involved in the initiation and propagation of several inflammatory conditions, mainly type I hypersensitivity. We report an unanticipated role of fullerenes as a negative regulator of allergic mediator release that suppresses Ag-driven type I hypersensitivity. Human MC and peripheral blood basophils exhibited a significant inhibition of IgE dependent mediator release when preincubated with C(60) fullerenes. Protein microarray demonstrated that inhibition of mediator release involves profound reductions in the activation of signaling molecules involved in mediator release and oxidative stress. Follow-up studies demonstrated that the tyrosine phosphorylation of Syk was dramatically inhibited in Ag-challenged cells first incubated with fullerenes. In addition, fullerene preincubation significantly inhibited IgE-induced elevation in cytoplasmic reactive oxygen species levels. Furthermore, fullerenes prevented the in vivo release of histamine and drop in core body temperature in vivo using a MC-dependent model of anaphylaxis. These findings identify a new biological function for fullerenes and may represent a novel way to control MC-dependent diseases including asthma, inflammatory arthritis, heart disease, and multiple sclerosis.     

Antioxidant activity of supramolecular water-soluble fullerenes evaluated by beta-carotene bleaching assay

We investigated the antioxidant activity of supramolecular water-soluble fullerenes, polyvinylpyrrolidone (PVP)-entrapped C(60), and gamma-cyclodextrin (CD)-bi-capped C(60), based on comparable beta-carotene bleaching assay. Antioxidant activity against reactive oxygen species (ROS) generated by three different methods, (i) autoxidation of linoleic acid, (ii) hydrogen peroxide promoter, and (iii) photoirradiation, was evaluated as percent of inhibition relative to a control experiment in view of the bleaching rate constant (k(obs)) as well as the persistent absorbency of beta-carotene. Water-soluble fullerenes exhibit significant inhibitory effects on the oxidative discoloration of beta-carotene in any system.     

Coupling biomolecules to fullerenes through a molecular adapter

A novel biotinylated fullerene has been synthesized to facilitate the attachment of biotin-conjugated proteins to C(60) through the use of streptavidin as a molecular adapter. The strong biotin-streptavidin interaction enables the attachment of fullerenes to streptavidin and, because of the availability of four biotin-binding sites on streptavidin, to biotinylated biomolecules. The feasibility of this approach is demonstrated by using biotinylated alkaline phosphatase. Due to the insolubility of fullerenes in aqueous media, the immobilized enzyme can be eventually recovered by simple centrifugation with no significant loss in activity.     

Interaction of c(60)-fullerene and carboxyfullerene with proteins: docking and binding site alignment

The unique properties of fullerenes have raised the interest of using them for biomedical applications. Within this framework, the interactions of fullerenes with proteins have been an exciting research target, yet little is known about how native proteins can bind fullerenes, and what is the nature of these interactions. Moreover, though some proteins have been shown to interact with fullerenes, up to date, no crystal structure of such complexes was obtained. Here we report docking studies aimed at examining the interactions of fullerene in two forms (C60 nonsubstituted fullerene and carboxyfullerene) with four proteins that are known to bind fullerene derivatives: HIV protease, fullerene-specific antibody, human serum albumin, and bovine serum albumin. Our work provides docking models with detailed binding pockets information, which closely match available experimental data. We further compare the predicted binding sites using a novel multiple binding site alignment method. A high similarity between the physicochemical properties and surface geometry was found for fullerene's binding sites of HIV protease and the human and bovine serum albumins.     

Recent progresses in application of fullerenes in cosmetics

Cosmetic industry is a fast growing industry with the continuous development of new active ingredients for skin care products. Fullerene C(60) and its derivates have been subject of intensive research in the last few years. Fullerenes display a wide range of different biological activities. Strong antioxidant capacities and effective quenching radical oxygen species (ROS) made fullerenes suitable active compounds in the formulation of skin care products. Published evidence on biological activities of fullerenes relevant for their application in cosmetics use and examples of published patents are presented. Recent trends in the use of fullerenes in topical formulations and patents are reviewed. Future investigations covering application of fullerenes in skin care are discussed.     

Antioxidant action of sugar-pendant C60 fullerenes

The action of C₆₀ fullerene and its derivatives as a radical-scavenging antioxidant has received much attention, but their reactivity toward free radicals and antioxidant capacity have not been well elucidated yet. In the present study, the reactivity of the two types of water-soluble, sugar-pendant C₆₀ fullerenes, C₆₀-1S and C₆₀-2S, toward peroxyl radical and their effect against human plasma lipid peroxidation were measured. The rate constants for the reaction of C₆₀-1S and C₆₀-2S with peroxyl radicals were obtained from their effect on the bleaching of β-carotene in lipid-SDS micelle system as 4.6 × 10³ and 8.0 × 10³ M^?1 s^?1 at 37 °C, respectively. They inhibited the free radical-induced lipid peroxidation in human plasma in a concentration-dependent manner. These results suggest that the sugar-pendant fullerenes C₆₀-1S and C₆₀-2S act as a radical-scavenging antioxidant with the activity similar to the phenolic antioxidants.