title: Genotoxicity of unmodified and organo-modified montmorillonite.
abstract: The natural clay mineral montmorillonite (Cloisite) Na+) and an organo-modified montmorillonite (Cloisite 30B) were investigated for genotoxic potential as crude suspensions and as suspensions filtrated through a 0.2-microm pore-size filter to remove particles above the nanometre range. Filtered and unfiltered water suspensions of both clays did not induce mutations in the Salmonella/microsome assay at concentrations up to 141microg/ml of the crude clay, using the tester strains TA98 and TA100. Filtered and unfiltered Cloisite) Na+ suspensions in culture medium did not induce DNA strand-breaks in Caco-2 cells after 24h of exposure, as tested in the alkaline comet assay. However, both the filtered and the unfiltered samples of Cloisite 30B induced DNA strand-breaks in a concentration-dependent manner and the two highest test concentrations produced statistically significantly different results from those seen with control samples (p<0.01 and p<0.001) and (p<0.05 and p<0.01), respectively. The unfiltered samples were tested up to concentrations of 170microg/ml and the filtered samples up to 216microg/ml before filtration. When tested in the same concentration range as used in the comet assay, none of the clays produced ROS in a cell-free test system (the DCFH-DA assay). Inductively coupled plasma mass-spectrometry (ICP-MS) was used to detect clay particles in the filtered samples using aluminium as a tracer element characteristic to clay. The results indicated that clay particles were absent in the filtered samples, which was independently confirmed by dynamic light-scattering measurements. Detection and identification of free quaternary ammonium modifier in the filtered sample was carried out by HPLC-Q-TOF/MS and revealed a total concentration of a mixture of quaternary ammonium analogues of 1.57microg/ml. These findings suggest that the genotoxicity of organo-modified montmorillonite was caused by the organo-modifier. The detected organo-modifier mixture was synthesized and comet-assay results showed that the genotoxic potency of this synthesized organo-modifier was in the same order of magnitude at equimolar concentrations of organo-modifier in filtrated Cloisite) 30B suspensions, and could therefore at least partly explain the genotoxic effect of Cloisite) 30B.
authors.affiliation: Technical University of Denmark, National Food Institute, Mørkhøj Bygade, 19-2860 Søborg, Denmark.
authors.forename: Anoop Kumar
authors.initials: AK
authors.lastname: Sharma
authors.forename: Bjørn
authors.initials: B
authors.lastname: Schmidt
authors.forename: Henrik
authors.initials: H
authors.lastname: Frandsen
authors.forename: Nicklas Raun
authors.initials: NR
authors.lastname: Jacobsen
authors.forename: Erik Husfeldt
authors.initials: EH
authors.lastname: Larsen
authors.forename: Mona-Lise
authors.initials: ML
authors.lastname: Binderup
mesh_terms: D000535:Aluminum / Q000737:chemistry; D000818:Animals; D001546:Bentonite / Q000633:toxicity*; D018938:Caco-2 Cells; D020552:Comet Assay; D004249:DNA Damage; D006801:Humans; D008297:Male; D009152:Mutagenicity Tests; D009153:Mutagens / Q000633:toxicity*; D009930:Organic Chemicals / Q000633:toxicity*; D051381:Rats; D017208:Rats, Wistar; D017382:Reactive Oxygen Species / Q000378:metabolism; D012475:Salmonella / Q000235:genetics
chemical_list: D009153:Mutagens; D009930:Organic Chemicals; D017382:Reactive Oxygen Species; D001546:Bentonite; D000535:Aluminum