Genotoxicity of unmodified and organo-modified montmorillonite.
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.
Sharma
Anoop Kumar
AK
Technical University of Denmark, National Food Institute, Mørkhøj Bygade, 19-2860 Søborg, Denmark.
Schmidt
Bjørn
B
Frandsen
Henrik
H
Jacobsen
Nicklas Raun
NR
Larsen
Erik Husfeldt
EH
Binderup
Mona-Lise
ML
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
D009153:Mutagens; D009930:Organic Chemicals; D017382:Reactive Oxygen Species; D001546:Bentonite; D000535:Aluminum