Attributing effects of aqueous C60 nano-aggregates to tetrahydrofuran decomposition products in larval zebrafish by assessment of gene expression

BACKGROUND: C-60 is a highly insoluble nanoparticle that can form colloidal suspended aggregates in water, which may lead to environmental exposure in aquatic organisms. Previous research has indicated toxicity from C-60 aggregate; however, effects could be because of tetrahydrofuran (THF) vehicle used to prepare aggregates. OBJECTIVE: Our goal was to investigate changes in survival and gene expression in larval zebrafish Danio rerio after exposure to aggregates Of C-60 Prepared by two methods: a) stirring and sonication of C-60 in water (C-60-water); and b) suspension of C-60 in THF followed by rotovaping, resuspension in water, and sparging with nitrogen gas (THF-C-60) RESULTS: Survival of larval zebrafish was reduced in THF-C-60 and THF-water but not in C-60-water. The greatest differences in gene expression were observed in fish exposed to THF-C-60 and most (182) of these genes were similarly expressed in fish exposed to THF-water. Significant up-regulation (3- to 7-fold) of genes involved in controlling oxidative damage was observed after exposure to THF-C-60 and THF-water. Analyses of THF-C-60 and THF-water by gas chromatography-mass spectrometry did not detect THF but found THF oxidation products gamma-butyrolactone and tetrahydro-2-furanol. Toxicity of gamma-butyrolactone (72-hr lethal concentration predicted to kill 50% was 47 ppm) indicated effects in THF treatments can result from gamma-butyrolactone toxicity. CONCLUSION: This research is the first to link toxic effects directly to a THF degradation product (gamma-butyrolactone) rather than to C-60 and may explain toxicity attributed to C-60 in other investigations. The present work was first presented at the meeting "Overcoming Obstacles to Effective Research Design in Nanotoxicology" held 24-26 April 2006 in Cambridge, Massachusetts, USA.