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Antigenicity of fullerenes: antibodies specific for fullerenes and their characteristics.

The recent interest in using Buckminsterfullerene (fullerene) derivatives in biological systems raises the possibility of their assay by immunological procedures. This, in turn, leads to the question of the ability of these unprecedented polygonal structures, made up solely of carbon atoms, to induce the production of specific antibodies. Immunization of mice with a C60 fullerene derivative conjugated to bovine thyroglobulin yielded a population of fullerene-specific antibodies of the IgG isotype, showing that the immune repertoire was diverse enough to recognize and process fullerenes as protein conjugates. The population of antibodies included a subpopulation that crossreacted with a C70 fullerene as determined by immune precipitation and ELISA procedures. These assays were made possible by the synthesis of water-soluble fullerene derivatives, including bovine and rabbit serum albumin conjugates and derivatives of trilysine and pentalysine, all of which were characterized as to the extent of substitution and their UV-Vis spectra. Possible interactions of fullerenes with the combining sites of IgG are discussed based on the physical chemistry of fullerenes and previously described protein-fullerene interactions. They remain to be confirmed by the isolation of mAbs for x-ray crystallographic studies.

Animals

Fullerene lipids: synthesis of C60 fullerene derivatives bearing a long-chain saturated or unsaturated triester system.

Tris(hydroxymethyl)aminomethane was successfully esterified with saturated and unsaturated long-chain fatty acids. The resulting amino-triester intermediates were successively reacted with chloroacetyl chloride, sodium azide, and C60 fullerene. Spectral evidence showed that the aziridine ring is joined to the junction of 16,6]-fused rings of the fullerene. The structures of the various C60 fullerene derivatives bearing a long-chain saturated or unsaturated triester system were characterized by spectroscopic and spectrometric methods.

Acetates

Comparison of immunological effects of Fullerene C60 and raw soot from Fullerene production on alveolar macrophages and macrophage like cells in vitro.

Two carbon modifications, C60 and Raw Soot (RS) from fullerene production, were incubated with macrophages and macrophage-like cells. Their effects were compared both to DQ12 Quartz as a positive particle control known to damage BAM and HL60M, and LPS or zymosan as biological inducers of immunological reactions. C60 as well as RS are not cytotoxic as compared to DQ12. C60 seems to induce a lower chemotactic activity than RS and DQ12. Furthermore, C60 is of no or only very little influence on the formation of reactive oxygen species (ROS). RS itself is oxidatively active both with H2DFC-DA in in vitro cell culture and with KMB in a cell free incubation under the influence of light. It is not yet clear whether the in vitro effects of RS are caused by ROS formation by the cells and/or the oxidative activity due to light as indicated from the results with KMB.

Acetylglucosaminidase

In vivo biological behavior of a water-miscible fullerene: 14C labeling, absorption, distribution, excretion and acute toxicity.

BACKGROUND: Water-soluble fullerenes have recently been shown to exhibit considerable in vitro biological activity including cytotoxicity, site-selective DNA cleavage and inhibition of HIV protease. To assess the potential of these compounds as drugs, studies on the in vivo behavior of fullerenes are needed. We therefore set out to synthesize a radiolabeled, water-soluble fullerene, in order to obtain data on the oral absorption, distribution and excretion of this class of compounds. RESULTS: We synthesized a 14C-labeled water-soluble [60]fullerene using dipolar trimethylenemethane, which undergoes cycloaddition to [60]fullerene. When administered orally to rats, this compound was not efficiently absorbed and was excreted primarily in the feces. When injected intravenously, however, it was distributed rapidly to various tissues, and most of the material was retained in the body after one week. The compound was also able to penetrate the blood-brain barrier. Acute toxicity of the water-miscible fullerene was found to be quite low. CONCLUSIONS: Although the water-soluble fullerenes (and possibly their simple metabolites) are not acutely toxic, they are retained in the body for long periods, raising concerns about chronic toxic effects. The fact that fullerenes distribute rapidly to many tissues suggests that they may eventually be useful to deliver highly polar drugs through membranes to a target tissue, however, and they may even have applications in the delivery of drugs to the brain. Recent advances in fullerene synthetic chemistry may also make it possible to control fullerene absorption/excretion profiles in the future.

Administration, Oral

Photoinduced electron transfer from synthetic chlorophyll analogue to fullerene C60 on carbon paste electrode. Preparation of a novel solar cell.

Upon a carbon paste electrode, fullerene C60 and successively methyl pyropheophorbide-a (chlorin) were casted to prepare a chlorin-fullerene modified carbon paste electrode (CFE). Photocurrents on the CFE were produced by irradiating of visible lights (> 510 nm) in an aqueous solution of 0.05 M ethylenediaminetetraacetic acid and 0.1 M Na2SO4 at pH 6.7. Larger anodic photocurrent was induced by the CFE than by the carbon paste electrodes modified with either the fullerene or the chlorin. In addition, the photocurrent of the CFE was dependent upon the amount of fullerene casted. The photocurrent action spectra of the CFE (at 300 mV vs. Ag/Ag+) showed that photoinduced electron transfer occurred from the excited state of the chlorin to the fullerene and/or from the chlorin to the photoexcited fullerene, and the electron of the fullerene anion radical produced was then shifted to the carbon paste. Upon irradiation of > 375 nm lights, the anodic photocurrent of the CFE was enhanced by increase in the illuminated light power and reached 0.03 mA cm-2 in the present system.

Carbon

Fullerenes C60 and C70 in flames.

The fullerenes C60 and C70 were first identified in carbon vapour produced by laser irradiation of graphite, and have recently been produced in macroscopic quantities by vaporization of graphite with resistive heating. It has also been suggested that fullerenes might be formed in sooting flames, and indeed all-carbon ions with mass/charge ratios suggestive of fullerenes have been detected in flames. These species were assumed to have the cage structures of fullerenes, but the mass spectroscopic evidence could not establish this conclusively. We have now collected samples of condensible compounds and soot from hydrocarbon combustion under a range of conditions, and analysed these using conventional techniques in an effort to detect fullerenes. Spectroscopic studies reveal the presence of C60 and C70 in yields and ratios that depend on temperature, pressure, carbon/oxygen ratio and residence time in the flame. Control of these conditions allows optimal yields of 3 g of fullerenes per kilogram of fuel carbon burned, and variation of the C70/C60 ratio over the range 0.26-5.7.

Carbon

Fullerenes: an extraterrestrial carbon carrier phase for noble gases.

In this work, we report on the discovery of naturally occurring fullerenes (C60 to C400) in the Allende and Murchison meteorites and some sediment samples from the 65 million-year-old Cretaceous/Tertiary boundary layer (KTB). Unlike the other pure forms of carbon (diamond and graphite), fullerenes are extractable in an organic solvent (e.g., toluene or 1,2,4-trichlorobenzene). The recognition of this unique property led to the detection and isolation of the higher fullerenes in the Kratschmer/Huffmann arc evaporated graphite soot and in the carbon material in the meteorite and impact deposits. By further exploiting the unique ability of the fullerene cage structure to encapsulate and retain noble gases, we have determined that both the Allende and Murchison fullerenes and the KTB fullerenes contain trapped noble gases with ratios that can only be described as extraterrestrial in origin.

Carbon

Effects of acute and subchronic exposure of topically applied fullerene extracts on the mouse skin.

The recent discovery that fullerenes (C60) can be produced in macroscopic quantities has sparked much interest in the chemistry of this unusual molecule. Concerns have also arisen about the potential carcinogenic effects of this molecule. We have addressed the potential acute and subchronic toxic effects of fullerenes applied in benzene on the mouse skin. The acute toxic effects measured in this study included epidermal DNA synthesis and the induction of ornithine decarboxylase activity in the epidermis. At the topical dose of fullerenes used in these studies (i.e., 200 micrograms), we found no effect on either DNA synthesis or ornithine decarboxylase activity over a 72 hour time course after treatment. The subchronic effects of the fullerenes as a mouse skin tumor promoter was assessed by repeatedly applying the chemical to the skin after initiation with the polycyclic aromatic hydrocarbon, 7,12-dimethylbenzanthracene (DMBA). Repeated administration of the fullerenes for up to 24 weeks post-initiation did not result in either benign or malignant skin tumor formation, whereas promotion with the phorbol ester, 12-O-tetradecanoyl-phorbol- 13-acetate (TPA) resulted in the formation of benign skin tumors. Our data indicate that fullerenes applied in benzene at a likely industrial exposure level do not cause acute toxic effects on the mouse skin epidermis.

9,10-Dimethyl-1,2-benzanthracene

C60 and C70 fullerene isomers generated in flames. Detection and verification by liquid chromatography/mass spectrometry analyses.

Fullerenes C60 and C70, generated by combustion, have been shown previously to be produced in controlled laminar flames accompanied by other compounds having fullerene-like characteristics. Analysis of these additional compounds by high-performance liquid chromatography, coupled on-line with mass spectrometry has identified them as isomers of the C60 and C70 fullerenes. The newly observed isomers have characteristic UV spectra and are thermally unstable, undergoing conversion to the more stable fullerenes with a half-life of about 1 h in boiling toluene (111 degrees C). Isomers of C60 and C70 fullerenes previously have been studied theoretically, but not observed experimentally. The flame-generated material also contains C60O and C70O compounds, as well as C76 and higher carbon clusters.

Carbon

Photoinactivation of vesicular stomatitis virus with fullerene conjugated with methoxy polyethylene glycol amine.

Fullerene is known to effectively produce mainly singlet oxygen by absorbing light energy, and therefore may be useful as a virucidal photosensitizer. This study was designed to investigate the virucidal activity of a water-soluble fullerene derivative, which is conjugated with methoxy polyethylene glycol amine to enhance its water solubility. Vesicular stomatitis virus (VSV) was inactivated upon illumination with this water-soluble fullerene, in a concentration- or dose-dependent manner. The titer of VSV was reduced by >5log 10 in the presence of 10 mg/ml (400 microM) fullerene derivative with 120 J/cm2 white light irradiation. VSV inactivation was inhibited by oxygen removal or by the addition of sodium azide, a known singlet oxygen scavenger. The substitution of H2O by D2O, which is known to prolong the lifetime of singlet oxygen, promoted the virucidal activity. These results indicate that singlet oxygen may play a major role in VSV photoinactivation by the water-soluble fullerene derivative. The concentration needed for virus inactivation is higher than that of other sensitizers such as methylene blue.

Antiviral Agents

[Determination of the genotoxicity of fullerene C60 and fullerol using the method of somatic mosaics on cells of Drosophila melanogaster wing and SOS-chromotest].

Genotoxicity of fullerene C60 was been determined in a prokaryotic in vitro test and in an eukaryotic in vivo system. The SOS chromotest of fullerene C60 in the Escherichia coli strain PQ37 revealed no genotoxicity either with or without activation of the rat liver homogenate. To perform the somatic mutation and recombination genotoxicity test (SMART) on somatic wing cells, Drosophila melanogaster larvae were grown on a standard medium with or without fullerene dope. No statistically significant differences were observed at the same fullerene concentrations in the SOS chromotest (0.45 micrograms/ml). Only at the highest possible fullerene concentration of 2.24 micrograms per 1 ml medium, a slight genotoxic effect was observed in wing cells. Fullerol demonstrates no mutagenic effect at a concentration of 2.46 mg/ml.

Animals

Fullerene data mining using bibliometrics and database tomography

Database tomography (DT) is a textual database analysis system consisting of two major components: (1) algorithms for extracting multiword phrase frequencies and phrase proximities (physical closeness of the multiword technical phrases) from any type of large textual database, to augment (2) interpretative capabilities of the expert human analyst. DT was used to derive technical intelligence from a fullerenes database derived from the Science Citation Index and the Engineering Compendex. Phrase frequency analysis by the technical domain experts provided the pervasive technical themes of the fullerenes database, and phrase proximity analysis provided the relationships among the pervasive technical themes. Bibliometric analysis of the fullerenes literature supplemented the DT results with author/journal/institution publication and citation data. Comparisons of fullerenes results with past analyses of similarly structured near-earth space, chemistry, hypersonic/supersonic flow, aircraft, and ship hydrodynamics databases are made. One important finding is that many of the normalized bibliometric distribution functions are extremely consistent across these diverse technical domains and could reasonably be expected to apply to broader chemical topics than fullerenes that span multiple structural classes. Finally, lessons learned about integrating the technical domain experts with the data mining tools are presented.

Journal Article

Direct preparation of radioactive fullerenes as a tracer for applications.

The C60 and C70 fullerenes were irradiated by high-energy gamma-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 with 11C or 13N were isolated and detected in the liquid phase by radiochromatography. It was found that (1) not only 11C or 13N 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 with 11C yields has led to high yields.

Carbon

Biological applications of fullerenes.

Thus far, development of applications of fullerenes in biology has been hampered by the poor water solubility of fullerenes. In spite of such concerns, fullerenes have proved useful for a wide variety of biological applications. As derivatized and underivatized fullerenes continue to become increasingly available, additional applications and further development of those discussed in this article will invariably follow.

Animals

Synthesis and characteristics of [60]fullerene polysiloxane stationary phase for capillary gas chromatography.

A new type of fullerene-containing polysiloxane was synthesized by reacting [60]fullerene with azidopropyl polysiloxane directly. The polysiloxanes have been used successfully as stationary phases for capillary gas chromatography. They displayed high column efficiency, wide operational temperature and high thermostability, and exhibited unique selectivity for many organic substances, such as alkanes, alcohols, ketones and aromatic compounds. The stationary phase was especially suitable for separation of high boiling-point compounds like polycyclic aromatic hydrocarbons and phthalic esters, etc. It was also found that some alcoholic or aromatic positional isomers could be well separated on the column. The influence of the fullerene content on the separation was also investigated.

Carbon

Pharmacokinetics of a water-soluble fullerene in rats.

Fullerenes are the recently discovered third allotropic form of carbon. The biological activities of these compounds are being studied for various purposes. The bis(monosuccinimide) derivative of p p'-bis(2-amino-ethyl)-diphenyl-C60 (MSAD-C60) is a water-soluble fullerene derivative. MSAD-C60 has been shown to have antiviral activity against human immunodeficiency virus types 1 and 2 in vitro and to have virucidal and anti-human immunodeficiency virus protease activities. Moreover, MSAD-C60 has been shown to be well tolerated in mice after intraperitoneal administration. The purpose of the present study was to develop a high-performance liquid chromatographic analytical methodology for MSAD-C60 and to characterize the preclinical pharmacokinetics of the compound in rats. Following intravenous administration of the fullerene derivative at a dose of 15 mg/kg of body weight, the concentrations of MSAD-C60 in plasma declined either bi- or triexponentially. The mean terminal-phase half-life of MSAD-C60 was 6.8 +/- 1.1 h (mean +/- standard deviation). Binding studies indicated that the compound is greater than 99% bound to plasma proteins. The average total clearance of the compound was 0.19 +/- 0.06 liter/h/kg. Urine samples obtained 24 h after intravenous administration did not contain detectable levels of the compound, indicating the absence of a significant renal clearance mechanism. The steady-state volume of distribution of MSAD-C60 averaged 2.1 +/- 0.8 liters/kg, indicating that the compound distributes into tissues. At a dose of 15 mg/kg, MSAD-C60 appeared to be well tolerated. However, a dose of 25 mg/kg resulted in shortness of breath and violent movement of the rats, followed by death within 5 min of dosing. Further controlled toxicity studies are needed to fully evaluate the toxicity of the compound.

Animals