Long Y. Chiang

Polyhydroxylated C60, fullerenols, as glutamate receptor antagonists and neuroprotective agents

Derivatives of C60 have been shown to be effective free radical scavengers. Hence, many of the biological functions of fullerene are believed to be due to their antioxidant properties. Here we present evidence to show that fullerenols, that are caged fullerene oxides, exert their neuroprotective functions by blocking glutamate receptors and lowering the intracellular calcium, [Ca2+]i. In neuronal cultures, fullerenols reduce glutamate‐induced neurotoxicity by about 80% at 50μM. No significant effect was observed on H2O2/Fe2+‐induced neurotoxicity under the same conditions. Fullerenols were found to inhibit glutamate receptor binding in a dose‐dependent manner inhibiting 50% of glutamate binding at 50 μM. Furthermore, AMPA receptors were found to be more sensitive to fullerenols than NMDA and KA receptors. On the other hand, GABAA receptors and taurine receptors were not significantly affected by fullerenols at the same concentrations used, suggesting that fullerenols inhibit primarily the glutamate receptors. In addition, fullerenols were also found to lower glutamate (Glu) receptor‐induced elevation of [Ca2+]i, suggesting that the underlying mechanism of neuronal protective function of fullerenols is likely due to its ability to block the glutamate receptors and to reduce the level of [Ca2+]i. J. Neurosci. Res. 62:600–607, 2000.

Efficient one-flask synthesis of water-soluble [60]fullerenols

Abstract C 60 molecule exhibited high reactivity towards the addition reaction of nitrogen dioxide radicals (·NO 2 ), which were generated by reaction of sodium nitrite with conc. HNO 3 . This chemical functionalization of C 60 resulted in polynitro fullerenes, C 60 (NO 2 ) n . Hydrolysis of polynitro fullerenes in aqueous NaOH gave the corresponding polyhydroxylated fullerene derivatives (fullerenols) in moderate overall yields. Results from mass spectra (FAB, negative ion) of fullerenols indicated their structure as C 60 derivatives with at least 16 hydroxy groups.

Acute and Subacute Toxicity Study of Water-Soluble Polyalkylsulfonated C60 in Rats

Polyalkylsulfonated C60, or FC4S, a highly water-soluble caged fullerene derivative, is believed to be a free radical remover or an antioxidant in biological systems. A 50 mg/ml aqueous solution was prepared as a master solution and administered to female Sprague-Dawley CD(Crl:CDR (SD)BR) rats in a single-dose acute toxicity study or a 12-day subacute toxicity study where rats were given the solution daily. In a study of the median lethal dose (LD50), no rats died after oral administration, and thus FC4S was considered to nontoxic if administered orally. In an LD50 intraperitoneal injection study, rats died within 30 hr after injection; the LD50 was determined to be approximately 600 mg per kilogram of body weight. Rats injected with the compound intraperitoneally or intravenously immediately eliminated the compound through the kidney; the kidney appeared to be the primary target organ. The compound induced a distinct lysosome-overload nephrosis, a phagolysosomal nephropathy characterized by a tinctorial difference between the outer cortex and the inner cortex and the medulla. The affected outer cortex showed a diffuse degeneration, with the presence of numerous large vacuoles and cytoplasmic aggregates in the tubular epithelium. The phagolysosomal nephropathy was detected in rats after acute exposure as well as in the surviving rats following 1 intraperitoneal injection of 500 mg/kg or intravenous injection of 100 mg/kg. Ultrastructural investigation revealed numerous membranous conglomerates characteristic of phagolysosomal and/or lysosomal inclusions in the cytoplasm of the renal tubular epithelium. These conglomerates were confined to the vacuole, electron-dense, and unevenly stained. They varied in size and shape and were fused or aggregated. Occasional phagolysosomes were also observed in the endothelial cells of the peritubular plexus. A preliminary study of microsomal enzyme activity analysis revealed a suppression effect of liver cytochrome P-450–dependent monooxygenase activities, including cytochrome P-450, cytochrome b5, and benzo(a)pyrene hydroxylase, but an increased level of kidney cytochrome P-450–dependent monooxygenase activities, including NADPH-cytochrome P-450 reductase. The significance of these enzyme alterations was not well determined. Further study is needed to clarify the correlation between the alterations of microsomal enzyme activity and the nephropathy of lysosomal overload-induced changes. These changes may serve as a biological marker in toxicity screening tests for this class of compound.

Polyhydroxylated C60, Fullerenol, a Novel Free-radical Trapper, Prevented Hydrogen Peroxide- and Cumene Hydroperoxide-elicited Changes in Rat Hippocampus In-vitro

The role of polyhydroxylated C60 (fullerenol), a novel free‐radical trapper, in prevention of hydrogen peroxide‐ and cumene hydroperoxide‐elicited damage was studied in hippocampal slices from the rat in‐vitro. The interactions of polyhydroxylated C60, adenosine and 6,7‐dinitroquinoxaline‐2,3‐dione (DNQX) were also compared.

Photodynamic therapy with fullerenes in vivo: reality or a dream?

Photodynamic therapy (PDT) employs the combination of nontoxic photosensitizers and visible light that is absorbed by the chromophore to produce long-lived triplet states that can carry out photochemistry in the presence of oxygen to kill cells. The closed carbon-cage structure found in fullerenes can act as a photosensitizer, especially when functionalized to impart water solubility. Although there are reports of the use of fullerenes to carry out light-mediated destruction of viruses, microorganisms and cancer cells in vitro, the use of fullerenes to mediate PDT of diseases such as cancer and infections in animal models is less well developed. It has recently been shown that fullerene PDT can be used to save the life of mice with wounds infected with pathogenic Gram-negative bacteria. Fullerene PDT has also been used to treat mouse models of various cancers including disseminated metastatic cancer in the peritoneal cavity. In vivo PDT with fullerenes represents a new application in nanomedicine.

Suppression of microsomal cytochrome P450-dependent monooxygenases and mitochondrial oxidative phosphorylation by fullerenol, a polyhydroxylated fullerene C60.

The acute toxicity of fullerenol-1 was determined using mice pretreated intraperitoneally (i.p.) with polyhydroxylated C60 derivatives. The LD50 value of fullerenol-1 was estimated to be 1.2 g/kg. Pretreatments with 0.5 and 1.0 g/kg fullerenol-1 decreased cytochromes P450 and b5 contents, and NADPH-cytochrome P450 reductase, benzo[a]pyrene hydroxylase, 7-ethoxycoumarin O-deethylase, aniline hydroxylase, and erythromycin N-demethylase activities in liver microsomes. Pretreatments with 0.01 and 0.1 g/kg fullerenol-1 had no effect on these monooxygenases. Additions of fullerenol-1 to mouse liver microsomes suppressed monooxygenases activities toward benzo[a]pyrene, 7-ethoxycoumarin, aniline, and erythromycin with IC50 values of 42, 94, 102 and 349 microM, respectively. Fullerenol-1 exhibited noncompetitive and mixed-type of inhibition in benzo[a]pyrene hydroxylation and 7-ethoxycoumarin O-deethylation, respectively. Additions of fullerenol-1 to rat liver mitochondria resulted in a dose-dependent inhibition of ADP-induced uncoupling and markedly inhibited mitochondrial Mg2+ -ATPase activity with an IC50 value of 7.1 microM. These results demonstrate that fullerenol-1 can suppress the levels of the microsomal enzymes in vivo and decrease the activities of P450-dependent monooxygenase and mitochondrial oxidative phosphorylation in vitro.

Free radical scavenging activity of water-soluble fullerenols

Water-soluble polyhydroxylated fullerene derivatives (fullerenols) show excellent efficiency in eliminating superoxide radicals (O2–˙), generated by xanthine and xanthine oxidase, thus revealing the potential use of these compounds as novel potent free radical scavengers in biological systems.

The possible mechanisms of the antiproliferative effect of fullerenol, polyhydroxylated C60, on vascular smooth muscle cells

1 The possible mechanisms of the antiproliferative effect of polyhydroxylated fullerene (fullerenol), a novel free radical trapper, were studied in rat vascular smooth muscle cells (A7r5 cells) and compared with the effect of ascorbic acid. 2 Fullerenol‐1 and ascorbic acid inhibited the proliferative responses in a number of cells, including rat aortic smooth muscle cells (A7r5 cells), human coronary artery smooth muscle cells, and human CEM lymphocytes (CEM cells) in a concentration dependent manner. 3 At the concentration range of 10−6 to 10−2 M, fullerenol‐1 and ascorbic acid concentration‐dependently inhibited the proliferative responses stimulated by serum in A7r5 cells. Fullerenol‐1 was more potent than ascorbic acid. 4 The production of O2− induced by alloxan, a diabetogenic compound, was reduced by fullerenol‐1 (10−4 M) in the presence of A7r5 cells. 5 The cytosolic protein kinase C activity of A7r5 cells stimulated by phorbol ester was reduced by 10−3 M fullerenol‐1, but not ascorbic acid (10−4–10−2 M) and fullerenol‐1 at lower concentrations (10−6–10−4 M). 6 In contrast, the membraneous protein tyrosine kinase activity of A7r5 cells stimulated by foetal calf serum was significantly reduced by fullerenol‐1 (10−6–10−3 M) and ascorbic acid (10−4–10−2 M). Again, the inhibitory activity of fullerenol‐1 was greater than that of ascorbic acid. 7 Our results demonstrate that fullerenol‐1 and ascorbic acid exhibit inhibitory effects on transduction signals in addition to their antioxidative property. It is suggested that the antiproliferative effect of fullerenol‐1 on vascular smooth muscle cells may partly be mediated through the inhibition of protein tyrosine kinase.

Multi-hydroxy additions onto C60 fullerene molecules

An efficient aqueous acid chemistry for the preparation of fullerols, consisting of 14–15 hydroxy moieties in an average structure, from C60 molecules is described.

Shining light on nanotechnology to help repair and regeneration.

Phototherapy can be used in two completely different but complementary therapeutic applications. While low level laser (or light) therapy (LLLT) uses red or near-infrared light alone to reduce inflammation, pain and stimulate tissue repair and regeneration, photodynamic therapy (PDT) uses the combination of light plus non-toxic dyes (called photosensitizers) to produce reactive oxygen species that can kill infectious microorganisms and cancer cells or destroy unwanted tissue (neo-vascularization in the choroid, atherosclerotic plaques in the arteries). The recent development of nanotechnology applied to medicine (nanomedicine) has opened a new front of advancement in the field of phototherapy and has provided hope for the development of nanoscale drug delivery platforms for effective killing of pathological cells and to promote repair and regeneration. Despite the well-known beneficial effects of phototherapy and nanomaterials in producing the killing of unwanted cells and promoting repair and regeneration, there are few reports that combine all three elements i.e. phototherapy, nanotechnology and, tissue repair and regeneration. However, these areas in all possible binary combinations have been addressed by many workers. The present review aims at highlighting the combined multi-model applications of phototherapy, nanotechnology and, reparative and regeneration medicine and outlines current strategies, future applications and limitations of nanoscale-assisted phototherapy for the management of cancers, microbial infections and other diseases, and to promote tissue repair and regeneration.